Medical Biology - Cell Surface Specializations PDF

MED105 MEDICAL BIOLOGY SPECIALIZATIONS OF THE CELL SURFACE (CELL SURFACE MODIFICATIONS) Asst. Prof. Fulya Küçükcankurt Department of Medical Biology e-mail: [email protected] Levels of organization in a multicellular organism Tissues are organized into fo...

MED105 MEDICAL BIOLOGY SPECIALIZATIONS OF THE CELL SURFACE (CELL SURFACE MODIFICATIONS) Asst. Prof. Fulya Küçükcankurt Department of Medical Biology e-mail: [email protected] Levels of organization in a multicellular organism Tissues are organized into four broad categories based on structural and functional similarities Types of tissue: 1. Connective tissue 2. Epithelial tissue 3. Muscle tissue 4. Nervous tissue Epithelial cells are the building blocks of epithelial tissue  There are several different types of epithelial cells based on their shape and arrangement Surface epithelium is classified by: Number of layers Simple: one layer Pseudostratified: one layer, but appears to have several layers Stratified: multiple layers Shape of cells at surface Squamous: heightwidth Cell Surface Modifications Specializations of the cell surface, mediating various functions Provide transportation Provide movement Connecting the cells to each other 1)Apical domain specializations: Microvilli Cell Surface Modifications Stereocilia Cilia Flagellum 2)Lateral domain specializations: Cell-to-cell junctions: a)Occluding: tight junctions b)Anchoring: Zonulae adherentes Fasciae adherentes (only in cardiac muscle cells) Desmosomes c)Communicating: gap junctions 3)Basal domain specializations: Basement membrane Cell-to-extracellular matrix (ECM) junctions: Anchoring: Focal adhesions Hemidesmosomes  The cells make up ephitelium have 3 principal characteristics: Cell junctions Polarity Basement membrane Special characteristics of Ephitelial Cells Polarity:epithelial tissues always have an apical and basal surface Support by connective tissue:at the basal surface, both the epithelial tissue and the connective tissue contribute to the basement membrane Cellularity:cells are in close contact with each other with little or no intercellular space between them Junctional Complex:for both attachment and communication  Cell Polarity is a main feature of many types of cells Epithelial cells show polar differentiation They have apical, lateral and basal sufaces They exibit modifications on their surfaces  Apical surface is located on the side of the lumen, or external environment. This pole show apical membrane specializations which alter the shape of this surface Lateral surfaces is on the sides and typically allows for connections with neighboring cells. Basal surfaces is the bottom edge of the cell and is adjacent to the basal lamina of the extracellular matrix, which separates the epithelial cell from the surrounding connective tissue. Cell form, structure, and function variations inside a cell are referred to as cell polarity. APICAL SURFACE MODIFICATIONS The surface of the most cells have extensions They are used in cell movement, absorbtion. Three types of specializations Microvilli Cilia, flagella Stereocilia  I- SPECIALIZATIONS OF FREE SURFACE (APICAL SURFACE ) 1-Microvilli surface extensions that increase surface area Specialized for absorption (15-40x in intestinal tract, kidneys); sensory (taste buds, inner ear) Brush border= dense ‘fringe’ (mucus membrane) 2-Cilia, flagella hairlike projections, nonmotile (more common) & motile (respiratory tract & uterine tubes) Beat in waves to sweep mucus, oocyte, embryo Beat in cell surface 3-Stereocilia whiplike structure much longer than cilia tail of sperm  Transmission electron micrograph of microvilli at the apical 1-Microvilli surfaces of proximal tubule epithelial cells. Encyclopedic Reference of Genomics and Proteomics in Molecular Medicine pp 1116–1121 Finger-like projections from the cell surface (size: 0.1 μm width and 1–3 μm long) Functions: 1. Increase the surface area for diffusion and minimize any increase in volume 2. Cellular adhesion 3. Mechanotransduction 4. Absorption 5. Secretion The specific localization on microvilli of important functional membrane proteins such as glucose transporters, ion channels, ion pumps, and ion exchangers indicate the importance and diversity of microvillar functions Scanning electron microscopy micrograph showing microvilli that protrude from the cell surface of resting peripheral blood human T cells. Frontiers in Immunology-2020 Microvillus Structure Microvilli are made up of 5 main proteins: actin, fimbrin, villin, myosin (Myo1A), calmodulin Actin filaments are cross linked into closely packed bundles by actin bundling proteins; fimbrin villin Actin filaments are attached to the plasma membrane by lateral arms consisting of myosin I calmodulin Animation:Microvilli 2-Cilia and Flagella Cilia and flagella are microtubule-based projections of the plasma membrane Cilia act as antennae that sense a variety of extracellular signals as well as being responsible for movement. There are 2 types of eukaryotic cilia, called primary (non- motile) cilia and motile cilia. Primary (non-motile) cilia are found on most animal cells and are involved in sensing extracellular signals, including movement, odorants, and light. Motile cilia are responsible for cell movement Cilia Light microscope Scanning electron microscope SEM  Cilia Classification 1)Motile cilia (flagella): Mechanical function. It is also found in single-celled organisms. In humans: - Airways cleaning - Sperm motility Motile cilia in respiratory tract - Conduction of the ovum from the fallopian tubes to the uterus - Ependymal cells in the choroid plexus located in the ventricles of the brain (CSF circulation!) 2)Non-motile (primary): They exist in multicellular organisms and function as "sensors". -They are found in almost every cell in humans. Primary cilia arising from a neuron  Motile Cilia 1-Surface of epithelial cells of the upper respiratory tract 2- Epithelial cells of the uterine tubes (oviducts ) 3- Epithelial cells of the efferent ducts (Ductus efferentes) The efferent ducts connect the testis with the epididymis. Uterine tubes (oviduct) Efferent ducts Respiratory tract Motile Cilia Functions 1-To move mucus over epithelial surfaces in respiratory epithelium toward the mouth. (Clear the mucus together with dust particles, dead cells and bacteria from respiratory passages) 2- In uterine tubes to transport the ovum toward the uterus. 3- In ductus efferentes (efferent ducts) to propel the spermatozoa toward the epididymis Uterine tubes (oviduct) Efferent ducts Respiratory tract The ciliated cells of the fallopian tube play a major role: transport of the ovum, the sperm cells and the zygote (a fertilized ovum)  Cells 2022, 11(9), 1416 The ciliated cells are located across the apical surface and facilitate the movement of mucus across the airway tract. Cilia cells : Move back and forth, carrying mucus up and out of the respiratory tract Non-Motile (Primer) Cilia Functions Developmental signaling pathways Wnt, Hedhehog Growth and differentiation Transforming growth factor β Platelet derived growth factor Sensory Photoreceptors, olfactory receptors Hormonal regulation MChr1 (Melanin-concentrating hormone receptor 1), 5HTr6 (5-Hydroxytryptamine (Serotonin) Receptor 6) Mechanical detection PCD1, PCD2 Two different sensory receptor cells have active transport systems served by their primary cilia Nature Reviews Genetics 6, 928–940 Basic Cilia Structure 1-Axoneme 2-Cell membrane 3-IFT (Intraflagellar Transport) 4-Basal body (derived from Centriole) 5- Cross section of the cilium 6- Microtubule triplets in basal body  Structures of primary (non-motile) and motile cilia Both non-motile and motile cilia are anchored in a centriole called a basal body, which contains nine triplets of microtubules 9+2 axoneme 9+0 axoneme Two of the microtubules in each triplet of the basal body are extended to form the axoneme. Diagram of ciliary structure Nature 448, 638–641 In each doublet ; Microtubule A is complete, it consist of 13 protofilaments in its wall (it has «O» shaped cross section) Microtubule B is incomplete, it consist of 10- 11 protofilament, it has a C shaped cross section. It is fused to Microtubule A and closes the defect in its Wall. Dynein arms Arranged along the length of the microtubule They are formed by a protein called dynein and contain ATPase (ATP splitting enzyme) activity. Nexin links Nexin links attach each microtubule A to the microtubule B of the adjacent doublet. ▪ Nexin links are composed of an elastic material called “nexin’’ ▪Nexin is the interdoublet link protein responsible for the maintenance of the nine-fold configuration in cilia and flagella. ▪ Responsible for recovery stroke (backward movement) MECHANISM OF CILIARY MOVEMENT Sliding Microtubule Mechanism The bending of an axoneme (A) When axonemes are exposed to the proteolytic enzyme trypsin, the linkages holding neighboring doublet microtubules together are broken. In this case, the addition of ATP allows the motor action of the dynein heads to slide one pair of doublet microtubules against the other pair. (B) In an intact axoneme (such as in a sperm), sliding of the doublet microtubules is prevented by flexible protein links. The motor action therefore causes a bending motion, creating waves or beating motions, Movement of Motile Cilia and Flagella Ciliary and flagellar movements are generated by microtubule sliding with axonemal dynein motors. Cilia and flagella are Cytoskeleton made up of microtubules. Movement= nine paired microtubule sets of the axoneme slide against one another causing cilia and flagella to bend. The motor protein dynein is responsible for generating the force required for movement. Ciliopathy Defects in ciliary activity cause a number of diseases Motile Ciliopathy: Immotile cilia syndrome, primary ciliary dyskinesia 1- Male infertility Normal number of spermatozoa but no motility 2- Respiratory tract disorders -Bronchiectasis (chronic dilation of bronchi ) -Chronic sinusitis 3-Situs inversus (complete transposition (right to left reversal) of the thoracic and abdominal organs Heart being in right, liver being in left The “gold-standard” diagnostic test for PCD has been electron microscopic Axial CT image showing situs inversus ultrastructural analysis of respiratory cilia obtained by nasal scrape or bronchial The liver is normally on the right side of the body and the spleen on the left, they are brush biopsy. switched in this patient with situs inversus. KARTAGENER SYNDROME  Electron Microscopic observation of immotile sperm flagellum of patients with Kartagener’s syndrome: Showed the absence of dynein arms Electrone microscope observation of bronchial biopsies showed no dynein arms in ciliary axonemes Dynein is essential for motility of cilia and flagella Kartagener syndrome is a genetic disease There is a congenital defect in the synthesis of dynein 1. Normal 2. Absent inner and outer dynein 3. Absent outer dynein 4. Absent inner dynein Genetics in Medicine volume 11, pages473–487 FLAGELLA Flagellum propels sperm 1- Longer than cilia Longest flagella are those of mammalian sperm 2- Same internal structure with cilia (Axoneme 9+2 ) 3-Different type of movement (undulating wave type of movement) Mammalian spermatozoa and flagellum structure 4- Less in number (one or two in a single cell) 5- Mammalian spermium contains 9 additional dense fibers arround the axoneme (9+9+2) (protective function) Scanning electron micrograph of a human sperm contacting a hamster egg. Stereocilia 1. Non-motile apical cell modifications 2. Long and irregular microvilli 3. Have no microtubule containing internal structure 4. Contain actin, lack of axoneme Bechara Kachar,NIH Localisations: epididymis, vas deferense, sensory of the inner ear Stereo cilia are found in the male reproductive tract and are thought to facilitate absorption in the epididymis and vas deferens. Stereocilia are essential in hearing and balance in inner ear.  Sensory cells in the inner ear The apical side of the inner ear hair cell carries the highly organized, actin-filled stereocilia Mechanotransduction function The stereocilia are anchored in the actin-rich cuticular plate. The kinocilium is located lateral to the largest stereocilium and is formed from the basal body. Damage to these cells results in decreased hearing sensitivity and balancing Stereocilia in male reproductive system In epididiymis, pseudostratified epithelium whose cells contain non-motile stereocilia. These stereocilia absorb much of the excess fluid containing the spermatozoa. The vas deferens is lined by a pseudostratified columnar epithelium composed of columnar cells and basal cells. The luminal surface of the columnar cell is lined by stereocilia. Epididymis Vas deferense Cell Membrane II Asst. Prof. Yalda Hekmatshoar Email: [email protected] MEMBRANE PROTEINS Although the lipid bilayer provides the basic structure of biological membranes, the membrane proteins perform most of the membrane’s specific tasks and therefore give each type of cell membrane its characteristic functional properties. The amounts and types of proteins in a membrane are highly variable. 25% protein in myelin membrane (serve as electrical insulation) Mitochondria internal membrane: 75% protein in membrane (serve as ATP production) Typical cell membrane: 50% protein by mass. Since proteins are much larger than lipids, this percentage corresponds to about one protein molecule per every 50 to 100 molecules of lipid. Cell membrane & organelle membranes each have unique collections of proteins.  Fluid mosaic model In 1972, Jonathan Singer and Garth Nicolson proposed the fluid mosaic model of membrane structure In this model, membranes are viewed as two- dimensional fluids in which proteins are inserted into lipid bilayers, The Cell: A Molecular Approach. 2nd edition. Cooper GM. Membrane Proteins Can Be Associated with the Lipid Bilayer in Various Ways Ø TRANSMEMBRANE PROTEINS Single Pass Alpha Helix, polypeptide chain crosses lipid bilayer only once (i.e.GF receptors) Multiple Pass Alpha Helix, polypeptide chain crosses multiple times (i.e.G protein coupled receptors) Beta Sheet Barrel (found in the outer membrane of mitochondria, chloroplasts, and many bacteria, porin proteins ) Ø Some others are anchored to the cytosolic surface by an amphiphilic alpha helix  multiple α helices a single α helix a rolled-up β sheet (a β barrel).  4- anchored to the cytosolic surface by an amphipathic alpha helix Proteins are located entirely in the cytosol and are attached to the cytosolic monolayer of the lipid bilayer, either by an amphiphilic α helix exposed on the surface of the protein 5- protein anchored to membrane by a fatty acid chain by one or more covalently attached lipid chains. The lipid-linked proteins are made as soluble proteins in the cytosol and are subsequently anchored to the membrane by the covalent attachment of the lipid group.  6-protein attached to the lipid bilayer by a oligosaccharide that is bound to phosphotidyl inositol (GPI anchor ) Other membrane proteins are entirely exposed at the external cell surface, being attached to the lipid bilayer only by a covalent linkage (via a specific oligosaccharide) to a lipid anchor in the outer monolayer of the plasma membrane. 7 and 8 - Peripheral proteins ü Membrane-associated proteins do not extend into the hydrophobic interior of the lipid bilayer at all; they are instead bound to either face of the membrane by noncovalent interactions with other membrane proteins. ü Many of the proteins of this type can be released from the membrane by relatively gentle extraction procedures, such as exposure to solutions of very high or low ionic strength or of extreme pH, which interfere with protein– protein interactions but leave the lipid bilayer intact; these proteins are often referred to as peripheral membrane proteins. ü Transmembrane proteins and many proteins held in the bilayer by lipid groups or hydrophobic polypeptide regions that insert into the hydrophobic core of the lipid bilayer cannot be released in these ways.  Classes of Membrane Proteins 1. Integral membrane proteins are inserted into the lipid bilayer, 2. Peripheral proteins are bound to the membrane indirectly by protein-protein interactions.  Integral proteins Many integral proteins are transmembrane proteins, Span the lipid bilayer with portions exposed on both sides of the membrane, These proteins can be visualized in electron micrographs of plasma membranes prepared by the freeze-fracture technique. The membrane-spanning portions of transmembrane proteins are usually α helices of 20 to 25 hydrophobic amino acids that are inserted into the membrane of the endoplasmic reticulum during synthesis of the polypeptide chain, These proteins are then transported in membrane vesicles from the endoplasmic reticulum to the Golgi apparatus, and from there to the plasma membrane, Carbohydrate groups are added to the polypeptide chains in both the endoplasmic reticulum and Golgi apparatus, so most transmembrane proteins of the plasma membrane are glycoproteins with their oligosaccharides exposed on the surface of the cell. The Cell: A Molecular Approach. 2nd edition. Cooper GM.  Transmembrane Secretory glycoproteins protein Golgi apparatus Vesicle ER ER lumen Glycolipid Plasma membrane: Cytoplasmic face Transmembrane Extracellular face glycoprotein Secreted protein Membrane glycolipid Integral Proteins They are amphipathic molecules with their hydrophilic region facing the cytoplasm or extracellular fluid and the hydrophobic domain interacting with the phospholipid tails. Integral proteins perform diverse functions such as transferring molecules and signals across the cell membrane. The extracellular portions of these proteins are usually glycosylated, as are the peripheral membrane proteins bound to the external face of the membrane.  Peripheral membrane proteins Peripheral membrane proteins associate with phospholipid heads or hydrophilic domains of integral proteins by non-covalent interactions. Many peripheral proteins also participate in cell signaling cascades as they can easily detach from the membrane. Other peripheral proteins link the membrane with the cytoskeleton, providing structural support. MEMBRANE PROTEINS Various functions: 1. Transporters 2. Enzymes 3. Cell-surface receptors 4. Cell-surface identity markers 5. Cell-to-cell adhesion proteins 6. Attachments to the cytoskeleton  Transport proteins Ø Transport molecules into and out of cells Ø Transport ATPases use energy of ATP to transfer ions across membrane. Allow passage of hydrophilic substances across the membrane Some transport proteins, called channel proteins, have a hydrophilic channel that certain molecules or ions can use as a tunnel Channel proteins called aquaporins facilitate the passage of water Other transport proteins, called carrier proteins, bind to molecules and change shape to shuttle them across the membrane A transport protein is specific for the substance it moves Enzyme: ØVarious enzymes allow different chemical reactions (membrane-associated reactions) Receptors: ØReceive signals from hormones, growth factors and other chemicals, Transmit those signals to the cell interior Cell-surface identity markers ØServe as antigens Cell-to-cell adhesion proteins ØAct as anchors for cytoskeletal components and receptors for extracellular matrix components (İntegrin: fibronectin, laminin receptor) 17 Membrane Proteins can serve as antigens Cell surface antigens are important in cell-to-cell recognition. Surface Proteins can stimulate the production of antibodies. Live mouse cells inject rat recognizes as foreign react with surface proteins on the mouse cells. Cell surface antigens : In humans the surface proteins responsible for recognition of cells as self or foreign are called Major Histocompatibility Complex MHC HLA markers ( Human leucocyte antigens ).  MHC molecules : Class I MHC proteins (on the surface of all nucleated cells) Class II MHC proteins (on the surface of B lymphocytes macrophages) MHC proteins contain constant and variable regions: Constant regions are identical in all individuals of a species. Variable regions show significant structural differences. MHC proteins of each individual are different from others (except identical twins) Receiving organism's immune system recognizes the foreign cells and initiates an immune response and rejects them.  MEMBRANE CARBOHYDRATES All eukaryotic cells have carbohydrate on their surface They are located only on the noncytosolic (outer) surface. (Asymmetrical) Glycoproteins CELL COAT GLYCOCALIX Glycolipids Both glycoproteins and glycolipids are abundant in eukaryotic cell membranes are absent from the inner mitochondrial membrane and the chloroplast lamellae. E.M. Micrograph of the surface of a lymphocyte stained with ruthenium red. The dark area: carbonhydrate-rich layer surrounding the cell Functions of membrane Carbohydrates 1- Important in cellular contacts and adhesion (Maintaining the integrity of the tissue) transient cell-cell adhesion processes, 2- Acts as a barrier to penetration of large particles. 3- Protect the plasma membrane against low pH,the effect of bile salts (intestinal epithelium) Prevents damage to cell membranes by digestive enzymes. 4- Cell to cell recognition, cell to matrix recognition. 5- Some glycolipids also act as binding sites for substances taken up by cells. i.e. Peptide hormones and some molecules that act as cell poisons. 6-Responsible for the various human blood types (A,B,O antigens) 7-Cell surface CH markers of red blood cells responsible for the ABO blood groups. Blood group antigens Arrangement of the sugar chains (genetically determined) of an individual with type A blood differ from those of an individual with type B blood. Carbohydrate portion constitute the antigenic determinant. Transfused blood will be recognized as foreign (if their membrane glycoproteins and glycolipids contain different carbohydrate markers) à Will cause immun response If carbohydrate organization of DONOR'S CELLS and RECIPIENT'S CELLS Ø are the same à No immunological response A,B,O antigens are structurally related oligosaccharides linked to lipids or proteins Protein Movement Proteins can move through the layer of the membrane similar to the lipids Can’t flip from one side to the other Are able to diffuse laterally through the membrane, This lateral movement was first shown directly in an experiment reported by Larry Frye and Michael Edidin in 1970, which provided support for the fluid mosaic model. Frye and Edidin fused human and mouse cells in culture to produce human-mouse cell hybrids Ø They analyzed the distribution of proteins in the membranes of these hybrid cells using antibodies that specifically recognize proteins of human and mouse origin. Ø These antibodies were labeled with different fluorescent dyes, so the human and mouse proteins could be distinguished by fluorescence microscopy. Ø Immediately after fusion, human and mouse proteins were localized to different halves of the hybrid cells. However, after a brief period of incubation at 37°C, the human and mouse proteins were completely intermixed over the cell surface, indicating that they moved freely through the plasma membrane. v The mobility of membrane proteins is restricted by, Cell cortex attachment (cytoskeleton) Extracellular attachment Attachment to other cells By diffusion barriers Tight junction – continuous barrier between adjacent cells MED105 INTRODUCTION to BASIC SCIENCES Asst. Prof. Behiye ÖZTÜRK ŞEN [email protected]  Lecture Presentation INTRODUCTION TO ORGANIC CHEMISTRY Altınbas University Lecturer: Asst. Prof. Behiye ÖZTÜRK ŞEN [email protected] MED105 INTRODUCTION to BASIC SCIENCES In this Chapter; You will learn What Organic Chemistry is Life and the Chemistry of Carbon Compounds Atomic Structure Isotopes Valence Electrons  What is Organic Chemistry? Organic chemistry plays a role in all aspects of our lives, ✔ from the clothing we wear, ✔ to the pixels of our television and computer screens, ✔ to preservatives in food, ✔ to the inks that color the pages of this book. Altınbas University Lecturer: Asst. Prof. Behiye ÖZTÜRK ŞEN [email protected] MED105 INTRODUCTION to BASIC SCIENCES  What is Organic Chemistry? Indeed, organic chemistry provides ❖ the power to synthesize new drugs, ❖ to engineer molecules that can make computer processors run more quickly, ❖ to understand why grilled meat can cause cancer and how its effects can be combated, and to design ways ❖ to knock the calories out of sugar while still making food taste deliciously sweet. It can explain biochemical processes like aging, neural functioning, and cardiac arrest, and show how we can prolong and improve life. Altınbas University Lecturer: Asst. Prof. Behiye ÖZTÜRK ŞEN [email protected] MED105 INTRODUCTION to BASIC SCIENCES  1.Life and the Chemistry of Carbon Compounds Organic chemistry is the chemistry of compounds that contain the element carbon. If a compound does not contain the element carbon, it is said to be inorganic Why C is important? There are several reasons, the primary one being this: carbon compounds are central to the structure of living organisms and therefore to the existence of life on Earth. We exist because of carbon compounds Altınbas University Lecturer: Asst. Prof. Behiye ÖZTÜRK ŞEN [email protected] MED105 INTRODUCTION to BASIC SCIENCES  What is it about carbon that makes it the element that nature has chosen for living organisms? There are two important reasons: 1. CARBON atoms can form strong bonds to other carbon atoms to form rings and chains of carbon atoms, And 2. CARBON atoms can also form strong bonds to elements such as hydrogen, nitrogen, oxygen, and sulfur. Because of these bond-forming properties, carbon can be the basis for the huge diversity of compounds necessary for the emergence of living organisms Altınbas University Lecturer: Asst. Prof. Behiye ÖZTÜRK ŞEN [email protected] MED105 INTRODUCTION to BASIC SCIENCES  1.2 Atomic Structure The compounds we encounter in chemistry are made up of elements combined in different proportions. Elements are made up of atoms. An atom consists of a dense, positively charged nucleus containing protons and neutrons and a surrounding cloud of electrons. Structure of Atom Altınbas University Lecturer: Asst. Prof. Behiye ÖZTÜRK ŞEN [email protected] MED105 INTRODUCTION to BASIC SCIENCES  1.2 Atomic Structure Each element is distinguished by its atomic number (Z), a number equal to the number of protons in its nucleus. the atomic number = the number of electrons surrounding the nucleus. (Neutral Atom) Altınbas University Lecturer: Asst. Prof. Behiye ÖZTÜRK ŞEN [email protected] MED105 INTRODUCTION to BASIC SCIENCES  1.2A Isotopes Although all the nuclei of all atoms of the same element will have the same number of protons, some atoms of the same element may have different masses because they have different numbers of neutrons. Such atoms are called isotopes. 12C, 13C, and 14C (ISOTOPES of C) All C have 6 Protons; but Neutron numbers are 6,7 and 8 respectively Altınbas University Lecturer: Asst. Prof. Behiye ÖZTÜRK ŞEN [email protected] MED105 INTRODUCTION to BASIC SCIENCES  1.2B Valence Electrons The most important shell, called the valence shell, is the outermost shell because the electrons of this shell are the ones that an atom uses in making chemical bonds with other atoms to form compounds. How do we know how many electrons an atom has in its valence shell? We look at the periodic table. The number of electrons in the valence shell (called valence electrons) is equal to the group number of the atom. For example, carbon is in group IVA and carbon has four valence electrons; oxygen is in group VIA and oxygen has six valence electrons. The halogens of group VIIA all have seven electrons. Altınbas University Lecturer: Asst. Prof. Behiye ÖZTÜRK ŞEN [email protected] MED105 INTRODUCTION to BASIC SCIENCES  Exercise How many valence electrons does each of the following atoms have? a) Na b) Cl c) Si d) B e) Ne f) N Altınbas University Lecturer: Asst. Prof. Behiye ÖZTÜRK ŞEN [email protected] MED105 INTRODUCTION to BASIC SCIENCES  ✔ Everywhere on Earth, organisms make organic molecules comprised almost exclusively of carbon, hydrogen, nitrogen, and oxygen. Sometimes a few slightly more exotic atoms, such as halogens and sulfur, are present. Globally, these compounds aid in day-to-day functioning of these organisms and/or their survival against predators. ✔ Organic molecules include many different compounds with diverse properties. For example, chlorophy ll in green plants harnesses the energy of sunlight, while vitamin C in citrus trees protects them against oxidative stress. ✔ Other molecules include capsaicin, a compound synthesized by pepper plants that wards off insects and birds that might try to eat them and is responsible for the “hotness” that we taste when we bite into a pepper. ✔ They also include salicylic acid, a signaling hormone made by willow trees, and lovastatin, a material found in oyster mushrooms that protects the mushroom against bacterial attacks. Altınbas University Lecturer: Asst. Prof. Behiye ÖZTÜRK ŞEN [email protected] MED105 INTRODUCTION to BASIC SCIENCES  Analgesic. It can modulate pain when applied to the skin and is currently sold under the tradename Capzacin a painkiller as well as an anti-acne medication a drug to decrease levels of cholesterol in human blood Altınbas University Lecturer: Asst. Prof. Behiye ÖZTÜRK ŞEN [email protected] MED105 INTRODUCTION to BASIC SCIENCES Altınbas University Lecturer: Asst. Prof. Behiye ÖZTÜRK ŞEN [email protected] MED105 INTRODUCTION to BASIC SCIENCES Prokaryotic and Eukaryotic cells Assist. Prof. Yalda Hekmatshoar Email: [email protected]  Definition of a cell A cell is the smallest unit that is capable of performing life functions, All living things are made of cells Smallest living unit of structure and function of all organisms is the cell  Life functions Reproduction and heredity Growth and development Metabolism – Chemical and physical life processes Movement and/or irritability – Respond to internal/external stimuli –Self propulsion Cell support, protection and storage mechanisms –Cell walls, vacuoles, granules, inclusions Transport of nutrients and waste  From simple microscopy, it has long been clear that living organisms can be classified on the basis of cell structure into two groups: ✓Prokaryotes (Bacteria and Archaea) –Pro= Before –Karyon= Kernel= nucleus ✓No nucleus ✓Eukaryotes (Everything else) –Eu= True Prokaryotes Prokaryotic cells were here first and for billions of years were the only form of life on Earth. Have no distinct nuclear compartment to house their DNA, Most prokaryotic cells are small and simple in outward appearance, ✓Most prokaryotic cells are 0.5–5µm,much smaller than the 10–100 µm of many eukaryotic cells. They live mostly as independent individuals or in loosely organized communities, rather than as multicellular organisms.  They are typically spherical or rod-shaped and measure a few micrometers in linear dimension. Molecular Cell Biology Lodish 5th edition  They often have a tough protective coat, called a cell wall, beneath which a plasma membrane encloses a single cytoplasmic compartment containing DNA, RNA, proteins, and the many small molecules needed for life. In the electron microscope, this cell interior appears as a matrix of varying texture without any discernible organized internal structure Molecular Cell Biology Lodish 5th edition  Prokaryotic cells live in an enormous variety of ecological niches, and they are astonishingly varied in their biochemical capabilities—far more so than eukaryotic cells. on plants & animals in plants & animals in the soil in depths of the oceans in extreme cold in extreme hot in extreme salt on the living on the dead ❖ Organotrophic species can utilize virtually any type of organic molecule as food, from sugars and amino acids to hydrocarbons and methane gas. ❖ Phototrophic species harvest light energy in a variety of ways, some of them generating oxygen as a by-product, others not. Molecular Biology o the cell, six edition ❖Lithotrophic species can feed on a plain diet of inorganic nutrients, getting their carbon from CO2 , and relying on H2 S to fuel their energy needs or on H2 , or Fe2+ , or elemental sulfur, or any of a host of other chemicals that occur in the environment. Molecular Biology o the cell, six edition ❖Prokaryotes (cells without a distinct nucleus) are biochemically the most diverse organisms and include species that can obtain all their energy and nutrients from inorganic chemical sources, such as the reactive mixtures of minerals released at hydrothermal vents on the ocean floor—the sort of diet that may have nourished the first living cells 3.5 billion years ago. ❖ DNA sequence comparisons reveal the family relationships of living organisms and show that the prokaryotes fall into two groups that diverged early in the course of evolution: the bacteria (or eubacteria) and the archaea. ❖ Together with the eukaryotes (cells with a membrane-enclosed nucleus), these constitute the three primary branches of the tree of life. ❖Most bacteria and archaea are small unicellular organisms with compact genomes comprising 1000–6000 genes.  The two groups of prokaryotes are called ✓bacteria (or eubacteria) ✓archaea (or archaebacteria).  The living world today is considered to consist of three major divisions or domains : ✓ bacteria, ✓Archaea, ✓eukaryotes Molecular Biology o the cell, six edition Archaea ✓are often found inhabiting environments that we humans avoid, such as bogs, sewage treatment plants, ocean depths, salt brines, and hot acid springs, Although they are also widespread in less extreme and more homely environments, from soils and lakes to the stomachs of cattle.  In outward appearance they are not easily distinguished from bacteria. At a molecular level, archaea seem to resemble eukaryotes more closely in their machinery for handling genetic information (replication, transcription, and translation), bacteria more closely in their apparatus for metabolism and energy conversion. Molecular Cell Biology Lodish 5th edition Eukaryotes Eukaryotes keep their DNA in a distinct membrane-enclosed intracellular compartment called the nucleus. (The name is from the Greek, meaning “truly nucleated,” from the words eu, “well” or “truly,” and karyon, “kernel” or “nucleus.”) Plants, fungi, and animals are eukaryotes  Eukaryotic cells, ✓ Are bigger and more elaborate than prokaryotic cells, ✓ Their genomes are bigger and more elaborate, ✓ The greater size is accompanied by radical differences in cell structure and function. ✓ Many classes of eukaryotic cells form multicellular organisms that attain levels of complexity unmatched by any prokaryote. Because they are so complex, Keep their DNA in an internal compartment called the nucleus. The nuclear envelope, a double layer of membrane, surrounds the nucleus and separates the DNA from the cytoplasm. Molecular Cell Biology Lodish 5th edition ✓ Their cells are, typically, 10 times bigger in linear dimension and 1000 times larger in volume. ✓ They have an elaborate cytoskeleton— ❖a system of protein filaments crisscrossing the cytoplasm and forming, together with the many proteins that attach to them, a system of girders, ropes, and motors that gives the cell mechanical strength, controls its shape, and drives and guides its movements. ✓ And the nuclear envelope is only one part of a set of internal membranes, each structurally similar to the plasma membrane and enclosing different types of spaces inside the cell, many of them involved in digestion and secretion. Modern Eukaryotic Cells Evolved from a Symbiosis ✓ All cells contain (or at one time did contain) mitochondria. ✓ These small bodies in the cytoplasm, enclosed by a double layer of membrane, take up oxygen and harness energy from the oxidation of food molecules—such as sugars— to produce most of the ATP that powers the cell’s activities. ✓ Mitochondria are similar in size to small bacteria, and, like bacteria, they have their own genome in the form of a circular DNA molecule, their own ribosomes that differ from those elsewhere in the eukaryotic cell, and their own transfer RNAs. ✓ It is now generally accepted that mitochondria originated from free-living oxygen-metabolizing (aerobic) bacteria that were engulfed by an ancestral cell that could otherwise make no such use of oxygen (that is, was anaerobic). Escaping digestion, these bacteria evolved in symbiosis with the engulfing cell and its progeny, receiving shelter and nourishment in return for the power generation they performed for their hosts. This partnership between a primitive anaerobic predator cell and an aerobic bacterial cell is thought to have been established about 1.5 billion years ago, when the Earth’s atmosphere first became rich in oxygen. What prokaryotic and eukaryotic cells have in common They both –have DNA as their genetic material –are covered by a cell membrane –contain RNA –are made from the same basic chemicals Carbohydrates, proteins, nucleicacids, minerals, fatsandvitamins –have ribosomes –regulate the flow of the nutrients and wastes that enter and leave them _ have similar basic metabolism like photo synthesis and reproduction –require a supply of energy Molecular Cell Biology Lodish 5th edition Fractionation of cells and analyzing their molecules Assist. Prof. Yalda Hekmatshoar  I) Examination of the cell ISOLATING CELLS AND GROWING THEM IN CULTURE Although the organelles and large molecules in a cell can be visualized with microscopes, understanding how these components function requires a detailed biochemical analysis. Most biochemical procedures require that large numbers of cells be physically disrupted to gain access to their components. To obtain as much information as possible about the cells in a tissue, biologists have developed ways of dissociating cells from tissues and separating them according to type. Molecular Biology o the cell, six edition  The first step in isolating individual cells is to disrupt the extracellular matrix and cell –cell junctions that hold the cells together. For this purpose, a tissue sample is typically treated with ✓ Proteolytic enzymes (such as trypsin and collagenase) to digest proteins in the extracellular matrix ✓ Agents (such as ethylene diamine tetraacetic acid, or EDTA) that bind, or chelate, the Ca2+ on which cell–cell adhesion depends. The tissue can then be teased apart into single cells by gentle agitation.  Cells Can Be Grown in Culture Tissue culture began in 1907 with an experiment designed to settle a controversy in neurobiology. Today, cultures are more commonly made from suspensions of cells dissociated from tissues. Cells grown in culture provide a more homogeneous population of cells from which to extract material, and they are also much more convenient to work with in the laboratory. Thermo Scientific Laminar flow cabinet clean bench II) Fraction of cells and analyzing their moleculs ✓ Cells Can Be Separated into Their Component Fractions A method to take the cells apart, separate subcellular components, and isolate major organelles and other subcellular components from one another. To purify a protein, it must first be extracted from inside the cell. ❖Cells can be broken up in various ways, They can be subjected to osmotic shock or ultrasonic vibration, forced through a small orifice, Ground up in a blender. ✓These procedures break many of the membranes of the cell (including the plasma membrane and endoplasmic reticulum) into fragments that immediately reseal to form small closed vesicles. If carefully carried out, organelles such as nuclei, mitochondria, the Golgi apparatus, lysosomes, and peroxisomes largely intact. The suspension of cells is thereby reduced to a thick slurry (called a homogenate or extract ) that contains a variety of membrane-enclosed organelles, each with a distinctive size, charge, and density. ❖ The different components of the homogenate must be separated. Such cell fractionations became possible only after the commercial development in the early 1940s of an instrument known as the preparative ultracentrifuge, which rotates extracts of broken cells at high speeds. This treatment separates cell components by size and density: in general, the largest objects experience the largest centrifugal force and move the most rapidly. ✓ At relatively low speed, large components such as nuclei sediment to form a pellet at the bottom of the centrifuge tube; ✓ At slightly higher speed, a pellet of mitochondria is deposited; ✓ At even higher speeds and with longer periods of centrifugation, first the small closed vesicles and then the ribosomes can be collected. All of these fractions are impure, but many of the contaminants can be removed by resuspending the pellet and repeating the centrifugation procedure several times. Centrifugation steps referred to in the figure are: low speed: 1000 times gravity for 10 minutes medium speed: 20,000 times gravity for 20 minutes high speed: 80,000 times gravity for 1 hour very high speed: 150,000 times gravity for 3 hours Molecular Biology o the cell, six edition Molecular Biology o the cell, six edition Cell Extracts Provide Accessible Systems to Study Cell Functions ❖Studies of organelles and other large subcellular components isolated in the ultracentrifuge have contributed enormously to our understanding of the functions of different cell components. ✓ Experiments on mitochondria and chloroplasts purified by centrifugation, for example, demonstrated the central function of these organelles in converting energy into forms that the cell can use. Resealed vesicles formed from fragments of rough and smooth endoplasmic reticulum (microsomes) have been separated from each other and analyzed as functional models of these compartments of the intact cell. Cell extracts also provide, in principle, the starting material for the complete separation of all of the individual macromolecular components of the cell. We now consider how this separation is achieved, focusing on proteins. Purified Cell-free Systems Are Required for the Precise Dissection of Molecular Functions Purified cell-free systems provide a means of studying biological processes free from all of the complex side reactions that occur in a living cell. To make this possible, cell homogenates are fractionated with the aim of purifying each of the individual macromolecules that are needed to catalyze a biological process of interest. Although much remains to be done, a great deal of what we know today about the molecular biology of the cell has been discovered by studies in such cell-free systems. They have been used, for example, to decipher the molecular details of DNA replication and DNA transcription, RNA splicing, protein translation, muscle contraction, and particle transport along microtubules, and many other processes that occur in cells. ANALYZING PROTEINS Proteins perform most cellular processes: ✓ They catalyze metabolic reactions, ✓ Major structural elements of the cell. The great variety of protein structures and functions has stimulated the development of a multitude of techniques to study them. ✓ Chromatography ✓ SDS Polyacrylamide-Gel Electrophoresis ✓ Two-Dimensional Gel Electrophoresis Proteins Can Be Separated by Chromatography Proteins are most often fractionated by column chromatography , in which a mixture of proteins in solution is passed through a column containing a porous solid matrix. Different proteins are retarded to different extents by their interaction with the matrix, and they can be collected separately as they flow out of the bottom of the column. Molecular Biology o the cell, six edition  Ion-exchange columns are packed with small beads that carry either a positive or a negative charge, so that proteins are fractionated according to the arrangement of charges on their surface. Hydrophobic columns are packed with beads from which hydrophobic side chains protrude, selectively retarding proteins with exposed hydrophobic regions. Gel-filtration columns , which separate proteins according to their size, are packed with tiny porous beads: molecules that are small enough to enter the pores linger inside successive beads as they pass down the column, while larger molecules remain in the solution flowing between the beads and therefore move more rapidly, emerging from the column first.  Depending on the choice of matrix, Proteins can be separated according to, ✓ Their charge (ion-exchange chromatography ), ✓ Their hydrophobicity (hydrophobic chromatography ), ✓ Their size (gel-filtration chromatography ), ✓ Their ability to bind to particular small molecules or to other macromolecules (affinity chromatography). Molecular Biology o the cell, six edition Proteins Can Be Separated by SDS Polyacrylamide-Gel Electrophoresis Proteins usually possess a net positive or negative charge, depending on the mixture of charged amino acids they contain. An electric field applied to a solution containing a protein molecule causes the protein to migrate at a rate that depends on its net charge and on its size and shape. The most popular application of this property is SDS polyacrylamide-gel electrophoresis (SDS-PAGE). It uses a highly cross-linked gel of polyacrylamide as the inert matrix through which the proteins migrate. The gel is prepared by polymerization of monomers; the pore size of the gel can be adjusted so that it is small enough to retard the migration of the protein molecules of interest.  The proteins are dissolved in a solution that includes a powerful negatively charged detergent, sodium dodecyl sulfate, or SDS. ❖ Because this detergent binds to hydrophobic regions of the protein molecules, causing them to unfold into extended polypeptide chains, the individual protein molecules are released from their associations with other proteins or lipid molecules and rendered freely soluble in the detergent solution. ❖ SDS-PAGE is widely used because it can separate all types of proteins, including those that are normally insoluble in water—such as the many proteins in membranes. Reducing agent such as β-mercaptoethanol is usually added to break any S–S linkages in the proteins, so that all of the constituent polypeptides in multisubunit proteins can be analyzed separately.  This method separates polypeptides by size, it provides information about the molecular weight and the subunit composition of proteins. Molecular Biology o the cell, six edition Two-Dimensional Gel Electrophoresis Provides Greater Protein Separation Because different proteins can have similar sizes, shapes, masses, and overall charges, most separation techniques such as SDS polyacrylamide-gel electrophoresis or ion- exchange chromatography cannot typically separate all the proteins in a cell or even in an organelle. In contrast, two-dimensional gel electrophoresis, which combines two different separation procedures, can resolve up to 2000 proteins in the form of a two-dimensional protein map.  In the first step, the proteins are separated by their intrinsic charges. ✓ The sample is dissolved in a small volume of a solution containing a nonionic (uncharged) detergent, together with β-mercaptoethanol and the denaturing reagent urea. ✓ This solution solubilizes, denatures, and dissociates all the polypeptide chains but leaves their intrinsic charge unchanged. The polypeptide chains are then separated in a pH gradient by a procedure called isoelectric focusing, which takes advantage of the variation in the net charge on a protein molecule with the pH of its surrounding solution. Every protein has a characteristic isoelectric point, the pH at which the protein has no net charge and therefore does not migrate in an electric field. In isoelectric focusing, proteins are separated electrophoretically in a narrow tube of polyacrylamide gel in which a gradient of pH is established by a mixture of special buffers. Each protein moves to a position in the gradient that corresponds to its isoelectric point and remains there.  In the second step, the narrow tube gel containing the separated proteins is again subjected to electrophoresis but in a direction that is at a right angle to the direction used in the first step. This time SDS is added, and the proteins separate according to their size, as in one- dimensional SDS-PAGE: the original tube gel is soaked in SDS and then placed along the top edge of an SDS polyacrylamide-gel slab, through which each polypeptide chain migrates to form a discrete spot. Even trace amounts of each polypeptide chain can be detected on the gel by various staining procedures—or by autoradiography if the protein sample was initially labeled with a radioisotope. The technique has such great resolving power that it can distinguish between two proteins that differ in only a single charged amino acid, or a single negatively charged phosphorylation site. MED105 INTRODUCTION to BASIC SCIENCES Asst. Prof. Behiye ÖZTÜRK ŞEN [email protected]  Lecture Presentation Basic concepts Chemical bonds Altınbas University Lecturer: Asst. Prof. Behiye ÖZTÜRK ŞEN [email protected] MED101 MOLECULLAR AND CELLULAR MEDICAL SCIENCES I In this Chapter; You will learn Chemical Bonds: The Octet Rule Ionic Bonds Covalent Bonds and Lewis Structures How To Write Lewis Structures Exceptions to the Octet Rule Formal Charges and How To Calculate Them A Summary of Formal Charges How To Write and Interpret Structural Formulas More About Dash Structural Formulas Condensed Structural Formulas Rules for Writing Resonance Structures  Chemical Bonds: The Octet Rule 1. Ionic (or electrovalent) bonds are formed by the transfer of one or more electrons from one atom to another to create ions. 2. Covalent bonds result when atoms share electrons. The central idea in their work on bonding is that atoms without the electronic configuration of a noble gas generally react to produce such a configuration because these configurations are known to be highly stable. For all of the noble gases except helium, this means achieving an octet of electrons in the valence shell. The valence shell is the outermost shell of electrons in an atom. The tendency for an atom to achieve a configuration where its valence shell contains eight electrons is called the octet rule. Altınbas University Lecturer: Asst. Prof. Behiye ÖZTÜRK ŞEN [email protected] MED105 INTRODUCTION to BASIC SCIENCES  Ionic Bonds Atoms may gain or lose electrons and form charged particles called ions. An ionic bond is an attractive force between oppositely charged ions. One source of such ions is a reaction between atoms of widely differing electronegativities Electronegativity is a measure of the ability of an atom to attract electrons. Altınbas University Lecturer: Asst. Prof. Behiye ÖZTÜRK ŞEN [email protected] MED105 INTRODUCTION to BASIC SCIENCES  Exercise Altınbas University Lecturer: Asst. Prof. Behiye ÖZTÜRK ŞEN [email protected] MED105 INTRODUCTION to BASIC SCIENCES  Covalent Bonds and Lewis Structures Covalent bonds form by sharing of electrons between atoms of similar electronegativities to achieve the configuration of a noble gas. Molecules are composed of atoms joined exclusively or predominantly by covalent bonds. Molecules may be represented by electron-dot formulas or, more conveniently, by formulas where each pair of electrons shared by two atoms is represented by a line. A dash structural formula has lines that show bonding electron pairs and includes elemental symbols for the atoms in a molecule. Altınbas University Lecturer: Asst. Prof. Behiye ÖZTÜRK ŞEN [email protected] MED105 INTRODUCTION to BASIC SCIENCES  Covalent Bonds and Lewis Structures 1. Hydrogen, being in group IA of the periodic table, has one valence electron. Two hydrogen atoms share electrons to form a hydrogen molecule, H 2. 2. Because chlorine is in group VIIA, its atoms have seven valence electrons. Two chlorine atoms can share electrons (one electron from each) to form a molecule of Cl 2. Altınbas University Lecturer: Asst. Prof. Behiye ÖZTÜRK ŞEN [email protected] MED105 INTRODUCTION to BASIC SCIENCES  Exercise Consider the following compounds and decide whether the bond in them would be IONIC Por COVALENT. (a) KCl (b) F2 (c) PH3 (d) CBr4 Altınbas University Lecturer: Asst. Prof. Behiye ÖZTÜRK ŞEN [email protected] MED105 INTRODUCTION to BASIC SCIENCES  How To Write Lewis Structures Several simple rules allow us to draw proper Lewis structures: 1. Lewis structures show the connections between atoms in a molecule or ion using only the valence electrons of the atoms involved. Valence electrons are those of an atom’s outermost shell. 2. For main group elements, the number of valence electrons a neutral atom brings to a Lewis structure is the same as its group number in the periodic table. Carbon, for example, is in group IVA and has four valence electrons; The halogens (e.g.,fluorine) are in group VIIA and each has seven valence electrons; hydrogen is in group IA and has one valence electron. Altınbas University Lecturer: Asst. Prof. Behiye ÖZTÜRK ŞEN [email protected] MED105 INTRODUCTION to BASIC SCIENCES  How To Write Lewis Structures 3. If the structure we are drawing is a negative ion (an anion), we add one electron for each negative charge to the original count of valence electrons. If the structure is a positive ion (a cation), we subtract one electron for each positive charge. Altınbas University Lecturer: Asst. Prof. Behiye ÖZTÜRK ŞEN [email protected] MED105 INTRODUCTION to BASIC SCIENCES  How To Write Lewis Structures 4. In drawing Lewis structures we try to give each atom the electron configuration of a noble gas. To do so, we draw structures where atoms share electrons to form covalent bonds or transfer electrons to form ions. a. Hydrogen forms one covalent bond by sharing its electron with an electron of another atom so that it can have two valence electrons, the same number as in the noble gas helium. b. Carbon forms four covalent bonds by sharing its four valence electrons with four valence electrons from other atoms, so that it can have eight electrons (the same as the electron configuration of neon, satisfying the octet rule). c. To achieve an octet of valence electrons, elements such as nitrogen, oxygen, and the halogens typically share only some of their valence electrons through covalent bonding, leaving others as unshared electron pairs. Nitrogen typically shares three electrons, oxygen two, and the halogens one. Altınbas University Lecturer: Asst. Prof. Behiye ÖZTÜRK ŞEN [email protected] MED105 INTRODUCTION to BASIC SCIENCES  Exercise Write the Lewis structure of CH3F. 1. We find the total number of valence electrons of all the atoms: Altınbas University Lecturer: Asst. Prof. Behiye ÖZTÜRK ŞEN [email protected] MED105 INTRODUCTION to BASIC SCIENCES  2. We use pairs of electrons to form bonds between all atoms that are bonded to each other. We represent these bonding pairs with lines. In our example this requires four pairs of electrons (8 of the 14 valence electrons). Altınbas University Lecturer: Asst. Prof. Behiye ÖZTÜRK ŞEN [email protected] MED105 INTRODUCTION to BASIC SCIENCES  3.We then add the remaining electrons in pairs so as to give each hydrogen 2 electrons (a duet) and every other atom 8 electrons (an octet). In our example, we assign the remaining 6 valence electrons to the fluorine atom in three nonbonding pairs. Altınbas University Lecturer: Asst. Prof. Behiye ÖZTÜRK ŞEN [email protected] MED105 INTRODUCTION to BASIC SCIENCES  Exercise Write a Lewis structure for methylamine (CH3NH2). 1. We find the total number of valence electrons for all the atoms. Altınbas University Lecturer: Asst. Prof. Behiye ÖZTÜRK ŞEN [email protected] MED105 INTRODUCTION to BASIC SCIENCES  2. We use one electron pair to join the carbon and nitrogen. Altınbas University Lecturer: Asst. Prof. Behiye ÖZTÜRK ŞEN [email protected] MED105 INTRODUCTION to BASIC SCIENCES  3. We use three pairs to form single bonds between the carbon and three hydrogen atoms. 4. We use two pairs to form single bonds between the nitrogen atom and two hydrogen atoms. 5. This leaves one electron pair, which we use as a lone pair on the nitrogen atom. Altınbas University Lecturer: Asst. Prof. Behiye ÖZTÜRK ŞEN [email protected] MED105 INTRODUCTION to BASIC SCIENCES  Exercise Write the Lewis structure of CH3OH. 5. If necessary, we use multiple bonds to satisfy the octet rule (i.e., give atoms the noble gas configuration). The carbonate ion (CO32−) illustrates this: Altınbas University Lecturer: Asst. Prof. Behiye ÖZTÜRK ŞEN [email protected] MED105 INTRODUCTION to BASIC SCIENCES  Exercise Write the Lewis structure of CH2O (formaldehyde). 1. Find the total number of valence electrons of all the atoms: Altınbas University Lecturer: Asst. Prof. Behiye ÖZTÜRK ŞEN [email protected] MED105 INTRODUCTION to BASIC SCIENCES  2. (a) Use pairs of electrons to form single bonds. Altınbas University Lecturer: Asst. Prof. Behiye ÖZTÜRK ŞEN [email protected] MED105 INTRODUCTION to BASIC SCIENCES  (b)Determine which atoms already have a full valence shell and which ones do not, and how many valence electrons we have used so far. In this case, we have used 6 valence electrons, and the valence shell is full for the hydrogen atoms but not for the carbon and oxygen atoms. (c) We use the remaining electrons as bonds or unshared electron pairs, to fill the valence shell of any atoms whose valence shell is not yet full, taking care not to exceed the octet rule. In this case 6 of the initial 12 valence electrons are left to use. We use 2 electrons to fill the valence shell of the carbon by another bond to the oxygen, and the remaining 4 electrons as two unshared electron pairs with the oxygen, filling its valence shell. Altınbas University Lecturer: Asst. Prof. Behiye ÖZTÜRK ŞEN [email protected] MED105 INTRODUCTION to BASIC SCIENCES  Exercise Write a Lewis structure for the toxic gas hydrogen cyanide (HCN). 1. We find the total number of valence electrons on all of the atoms: Altınbas University Lecturer: Asst. Prof. Behiye ÖZTÜRK ŞEN [email protected] MED105 INTRODUCTION to BASIC SCIENCES  2. We use one pair of electrons to form a single bond between the hydrogen atom and the carbon atom, and we use three pairs to form a triple bond between the carbon atom and the nitrogen atom. This leaves two electrons. We use these as an unshared pair on the nitrogen atom. Now each atom has the electronic structure of a noble gas. The hydrogen atom has two electrons (like helium) and the carbon and nitrogen atoms each have eight electrons (like neon). Altınbas University Lecturer: Asst. Prof. Behiye ÖZTÜRK ŞEN [email protected] MED105 INTRODUCTION to BASIC SCIENCES  Exceptions to the Octet Rule Elements of the second period of the periodic table can have a maximum of four bonds (i.e., have eight electrons around them) because these elements have only one 2s and three 2p orbitals available for bonding. Each orbital can contain two electrons, and a total of eight electrons fills these orbitals. The octet rule, therefore, only applies to these elements, and even here, as we shall see in compounds of beryllium and boron, fewer than eight electrons are possible. Elements of the third period and beyond have d orbitals that can be used for bonding. These elements can accommodate more than eight electrons in their valence shells and therefore can form more than four covalent bonds. Examples are compounds such as PCl5 and SF6. Altınbas University Lecturer: Asst. Prof. Behiye ÖZTÜRK ŞEN [email protected] MED105 INTRODUCTION to BASIC SCIENCES  Exceptions to the Octet Rule Altınbas University Lecturer: Asst. Prof. Behiye ÖZTÜRK ŞEN [email protected] MED105 INTRODUCTION to BASIC SCIENCES  Exercise Write a Lewis structure for the sulfate ion (SO42−) 1.We find the total number of valence electrons including the extra 2 electrons needed to give the ion the double negative charge: Altınbas University Lecturer: Asst. Prof. Behiye ÖZTÜRK ŞEN [email protected] MED105 INTRODUCTION to BASIC SCIENCES  2.We use four pairs of electrons to form bonds between the sulfur atom and the four oxygen atoms: Altınbas University Lecturer: Asst. Prof. Behiye ÖZTÜRK ŞEN [email protected] MED105 INTRODUCTION to BASIC SCIENCES  3.We add the remaining 24 electrons as unshared pairs on oxygen atoms and as double bonds between the sülfür atom and two oxygen atoms. This gives each oxygen 8 electrons and the sulfur atom 12: Altınbas University Lecturer: Asst. Prof. Behiye ÖZTÜRK ŞEN [email protected] MED105 INTRODUCTION to BASIC SCIENCES  Formal Charges and How To Calculate Them Many Lewis structures are incomplete until we decide whether any of their atoms have a formal charge. Calculating the formal charge on an atom in a Lewis structure is simply a bookkeeping method for its valence electrons. First, we examine each atom and, using the periodic table, we determine how many valence electrons it would have if it were an isolated atom. This is equal to the group number of the atom in the periodic table. For hydrogen this number equals 1, for carbon it equals 4, for nitrogen it equals 5, and for oxygen it equals 6. Next, we examine the atom in the Lewis structure and we assign the valence electrons in the following way: Altınbas University Lecturer: Asst. Prof. Behiye ÖZTÜRK ŞEN [email protected] MED105 INTRODUCTION to BASIC SCIENCES  Formal Charges and How To Calculate Them We assign to each atom half of the electrons it is sharing with another atom and all of its unshared (lone) electron pairs. Then we do the following calculation for the atom: Formal charge = number of valence electrons − 1/2 number of shared electrons − number of unshared electrons or F = Z − (1/2)S − U the number of unshared electrons formal charge group number of the element the number of shared electrons It is important to note, too, that the arithmetic sum of all the formal charges in a molecule or ion will equal the overall charge on the molecule or ion. Altınbas University Lecturer: Asst. Prof. Behiye ÖZTÜRK ŞEN [email protected] MED105 INTRODUCTION to BASIC SCIENCES  Exercise Formal Charges of Ammonium Ion or Altınbas University Lecturer: Asst. Prof. Behiye ÖZTÜRK ŞEN [email protected] MED105 INTRODUCTION to BASIC SCIENCES  Exercise Nitrate Ion or Altınbas University Lecturer: Asst. Prof. Behiye ÖZTÜRK ŞEN [email protected] MED105 INTRODUCTION to BASIC SCIENCES  Exercise Water or Altınbas University Lecturer: Asst. Prof. Behiye ÖZTÜRK ŞEN [email protected] MED105 INTRODUCTION to BASIC SCIENCES  Exercise Ammonia or Altınbas University Lecturer: Asst. Prof. Behiye ÖZTÜRK ŞEN [email protected] MED105 INTRODUCTION to BASIC SCIENCES  A Summary of Formal Charges Altınbas University Lecturer: Asst. Prof. Behiye ÖZTÜRK ŞEN [email protected] MED105 INTRODUCTION to BASIC SCIENCES  How To Write and Interpret Structural Formulas Altınbas University Lecturer: Asst. Prof. Behiye ÖZTÜRK ŞEN [email protected] MED105 INTRODUCTION to BASIC SCIENCES  More About Dash Structural Formulas Dash structural formulas have lines that show bonding electron pairs, and include elemental symbols for all of the atoms in a molecule.  Condensed Structural Formulas In condensed formulas all of the hydrogen atoms that are attached to a particular carbon are usually written immediately after the carbon Altınbas University Lecturer: Asst. Prof. Behiye ÖZTÜRK ŞEN [email protected] MED105 INTRODUCTION to BASIC SCIENCES  Condensed Structural Formulas The condensed formula for isopropyl alcohol can be written in four different ways: Altınbas University Lecturer: Asst. Prof. Behiye ÖZTÜRK ŞEN [email protected] MED105 INTRODUCTION to BASIC SCIENCES  Rules for Writing Resonance Structures 1. Resonance structures exist only on paper. Although they have no real existence of their own, resonance structures are useful because they allow us to describe molecules and ions for which a single Lewis structure is inadequate. We write two or more Lewis structures, calling them resonance structures or resonance contributors. We connect these structures by double-headed arrows (←→ ), and we say that the real molecule or ion is a hybrid of all of them. 2. We are only allowed to move electrons in writing resonance structures. The positions of the nuclei of the atoms must remain the same in all of the structures. Structure 3 is not a resonance structure of 1 or 2, for example, because in order to form it we would have to move a hydrogen atom and this is not permitted: Altınbas University Lecturer: Asst. Prof. Behiye ÖZTÜRK ŞEN [email protected] MED105 INTRODUCTION to BASIC SCIENCES  Rules for Writing Resonance Structures Altınbas University Lecturer: Asst. Prof. Behiye ÖZTÜRK ŞEN [email protected] MED105 INTRODUCTION to BASIC SCIENCES  3. All of the structures must be proper Lewis structures. We should not write structures in which carbon has five bonds, for example: Altınbas University Lecturer: Asst. Prof. Behiye ÖZTÜRK ŞEN [email protected] MED105 INTRODUCTION to BASIC SCIENCES  4. The energy of the resonance hybrid is lower than the energy of any contributing structure. Resonance stabilizes a molecule or ion. This is especially true when the resonance structures are equivalent. Chemists call this stabilization resonance stabilization. If the resonance structures are equivalent, then the resonance stabilization is large. Altınbas University Lecturer: Asst. Prof. Behiye ÖZTÜRK ŞEN [email protected] MED105 INTRODUCTION to BASIC SCIENCES Altınbas University Lecturer: Asst. Prof. Behiye ÖZTÜRK ŞEN [email protected] MED105 INTRODUCTION to BASIC SCIENCES  5. The more stable a structure is (when taken by itself ), the greater is its contribution to the hybrid. Altınbas University Lecturer: Asst. Prof. Behiye ÖZTÜRK ŞEN [email protected] MED105 INTRODUCTION to BASIC SCIENCES Cell Membrane I Asst. Prof. Yalda Hekmatshoar Email: [email protected]  Despite their differing functions, all biological membranes have a common general structure: veach is a very thin film of lipid and protein molecules, held together mainly by noncovalent interactions.  Cell membranes ü are crucial to the life of the cell. ü The plasma membrane encloses the cell, defines its boundaries, and maintains the essential differences between the cytosol and the extracellular environment. ü Inside eukaryotic cells, the membranes of the nucleus, endoplasmic reticulum, Golgi apparatus, mitochondria, and other membrane-enclosed organelles maintain the characteristic differences between the contents of each organelle and the cytosol. ü Ion gradients across membranes, established by the activities of specialized membrane proteins, can be used to synthesize ATP, to drive the transport of selected solutes across the membrane, or, as in nerve and muscle cells, to produce and transmit electrical signals. ü In all cells, the plasma membrane also contains proteins that act as sensors of external signals, allowing the cell to change its behavior in response to environmental cues, including signals from other cells; these protein sensors, or receptors, transfer information—rather than molecules— across the membrane.  Cell membranes are, This lipid bilayer provides the basic fluid structure of the membrane and serves as a relatively impermeable barrier to the passage of most water-soluble molecules. Most membrane proteins span the lipid bilayer and mediate nearly all of the other functions of the membrane, including the transport of specific molecules across it, and the catalysis of membrane-associated reactions such as ATP synthesis. In the plasma membrane, some transmembrane proteins serve as structural links that connect the cytoskeleton through the lipid bilayer to either the extracellular matrix or an adjacent cell, while others serve as receptors to detect and transduce chemical signals in the cell’s environment. It takes many kinds of membrane proteins to enable a cell to function and interact with its environment, and it is estimated that about 30% of the proteins encoded in an animal’s genome are membrane proteins. THE LIPID BILAYER ü Provides the basic structure for all cell membranes. ü It is easily seen by electron microscopy, and its bilayer structure is attributable exclusively to the special properties of the lipid molecules, which assemble spontaneously into bilayers even under simple artificial conditions. Phosphoglycerides, Sphingolipids, and Sterols Are the Major Lipids in Cell Membranes ü Lipid molecules constitute about 50% of the mass of most animal cell membranes, nearly all of the remainder being protein. ü All of the lipid molecules in cell membranes are amphiphilic —that is, they have a hydrophilic (“water-loving”) or polar end and a hydrophobic (“water-fearing”) or nonpolar end. Phospholipids ü The most abundant membrane lipids. ü These have a polar head group containing a phosphate group and two hydrophobic hydrocarbon tails. ü In animal, plant, and bacterial cells, the tails are usually fatty acids, and they can differ in length (they normally contain between 14 and 24 carbon atoms). One tail typically has one or more cis-double bonds (that is, it is unsaturated), while the other tail does not (that is, it is saturated). each cis-double bond creates a kink in the tail. Differences in the length and saturation of the fatty acid tails influence how phospholipid molecules pack against one another, thereby affecting the fluidity of the membrane. Phosphoglycerides üThe main phospholipids in most animal cell membranes, üwhich have a three-carbon glycerol backbone, üTwo long-chain fatty acids are linked through ester bonds to adjacent carbon atoms of the glycerol, and the third carbon atom of the glycerol is attached to a phosphate group, which in turn is linked to one of several types of head group. üBy combining several different fatty acids and head groups, cells make many different phosphoglycerides. üPhosphatidylethanolamine, phosphatidylserine, and phosphatidylcholine are the most abundant ones in mammalian cell membranes  sphingolipids ü Another important class of phospholipids ü They are built from sphingosine rather than glycerol, ü is a long acyl chain with an amino group (NH2) and two hydroxyl groups (OH) at one end. ü In sphingomyelin, the most common sphingolipid, a fatty acid tail is attached to the amino group, and a phosphocholine group is attached to the terminal hydroxyl group. ü Together, the phospholipids phosphatidylcholine, phosphatidylethanolamine, phosphatidylserine, and sphingomyelin constitute more than half the mass of lipid in most mammalian cell membranes.  In addition to phospholipids, the lipid bilayers in many cell membranes contain glycolipids and cholesterol. Glycolipids resemble sphingolipids, but, instead of a phosphate-linked head group, they have sugars attached. cholesterol ü Eukaryotic plasma membranes contain especially large amounts of cholesterol Ø up to one molecule for every phospholipid molecule. ü Cholesterol is a sterol. ü It contains a rigid ring structure, to which is attached a single polar hydroxyl group and a short nonpolar hydrocarbon chain. ü The cholesterol molecules orient themselves in the bilayer with their hydroxyl group close to the polar head groups of adjacent phospholipid molecules. Glycolipids üAre Found on the Surface of All Eukaryotic Plasma Membranes üSugar-containing lipid molecules called glycolipids ü have the most extreme asymmetry in their membrane distribution: these molecules, Øwhether in the plasma membrane or in intracellular membranes, are found exclusively in the monolayer facing away from the cytosol. Glycolipids probably occur in all eukaryotic cell plasma membranes, Øwhere they generally constitute about 5% of the lipid molecules in the outer monolayer. ü The shape and amphiphilic nature of the phospholipid molecules cause them to form bilayers spontaneously in aqueous environments. ü They spontaneously aggregate to bury their hydrophobic tails in the interior, where they are shielded from the water, and they expose their hydrophilic heads to water. ü Depending on their shape, they can do this in either of two ways: 1. They can form spherical micelles , with the tails inward, 2. They can form double-layered sheets, or bilayers , with the hydrophobic tails sandwiched between the hydrophilic head groups The Lipid Bilayer Is a Two-dimensional Fluid Around 1970, researchers first recognized that individual lipid molecules are able to diffuse freely within the plane of a lipid bilayer. The initial demonstration came from studies of synthetic (artificial) lipid bilayers, which can be made in the form of spherical vesicles, called liposomes ; or in the form of planar bilayers formed across a hole in a partition between two aqueous compartments or on a solid support.  Lipid asymmetry is functionally important, üEspecially in converting extracellular signals into intracellular ones. üMany cytosolic proteins bind to specific lipid head groups found in the cytosolic monolayer of the lipid bilayer. üThe enzyme protein kinase C (PKC), for example, which is activated in response to various extracellular signals, binds to the cytosolic face of the plasma membrane, where phosphatidylserine is concentrated, and requires this negatively charged phospholipid for its activity.  The translocation of the phosphatidylserine in apoptotic cells is thought to occur by two mechanisms: 1. The phospholipid translocator that normally transports this lipid from the outer monolayer to the inner monolayer is inactivated. 2. A “scramblase” that transfers phospholipids nonspecifically in both directions between the two monolayers is activated. MED105 MEDICAL BIOLOGY Department of Medical Biology Reference Books 1. The Cell – A Molecular Approach Geoffrey M. Cooper 2. Molecular Biology of the Cell B. Alberts, D Bray, J. Lewis, M. Raff, Keith Roberts, JD. Watson 3. Molecular Cell Biology J. Darrnell, H. Lodish, D Baltimore, Thomas D Pollard, WC Earnshaw 4. Medical Cell Biology Steven R. Goodman 5. Molecular and Cellular Biology Stephen L Wolfe 6. Human Molecular Biology RJ Epstein  MED 105 INTRODUCTION TO BASIC SCIENCES 1. Introduction to Medical Biology 2. Examination methods of cell biology; Cell cultures, hybrid cells 3. Fractionation of cells and analysis of their molecules 4. Prokaryotic and Eukaryotic cells 5. Structure and function of cellular membranes (1/2) 6. Structure and function of cellular membranes (2/2) 7. Modifications of the cell surface (1/2) 8. Modifications of the cell surface (2/2) 9. Cell-cell junctions 10. Cell-matrix junctions 11. Cell adhesion molecules 12. Cellular organelles;The mitochondrion 13. Structure, function and biosynthesis of ribosomes 14. Structure and function of the endoplasmic reticulum 15. Molecular mechanisms of protein synthesis in rough endoplasmic reticulum  MED 105 INTRODUCTION TO BASIC SCIENCES 16. The Golgi apparatus and protein sorting 17. Lysosome structure and function 18. Cytoskeleton (1/2): Microtubulles and intracellular transport 19. Cytoskeleton (2/2): Microfilaments and intermediate filaments 20. The nucleus, nuclear envelope, nuclear pores 21. Transport through the nuclear envelope 22. Nucleolus, nuclear matrix 23. Structure of Chromatin 24. Fine structure of chromatin 25. Global structure of chromosomes, centromere, telomeres Introduction to Cell Biology What is cell? CELLS are basic morphological and functional units of the body CELL THEORY: A Core Principle of Biology 1. All living organisms are composed of one of more cells 2. Cells are the fundamental units of both structure and function in all living organisms. 3. Cells arise only from preexisting cells with cells passing copies of their genetic material on the dougther cells  Magic of Life Although every living thing starts life with a single cell that looks "the same" Characteristics of living organisms Metabolism Response Homeostasis Growth Reproduction Excretion Nutrition  METHODS FOR EXAMINATION OF CELLS  How cells are studied I- Examination of II-Fractionation of the cell as a whole cells and analyzing (without disruption) their molecules A- Examination of Cells are disrupted living cells (Fresh and their organelles tissues) and macromolecules B- Examination of isolated in pure killed and preserved form tissues and cells I-Methods for examination of living cells (Vital examination ) Examination of living cells under a microscope is the oldest one of the techniques Looking at the structure of cells in Direct observation the microscope  Microscope A microscope is a scientific instrument that is used to magnify and observe objects that are too small to be seen with the naked eye. Microscopes are important tools in various scientific fields Light microscope Ø A light microscope is an optical instrument used to view objects too small to with the naked eye. Ø Eukaryotic organisms and bacteria are measured in micrometers, viruses in nanometers, atoms and molecules in angstroms. 1 mm = 1000 µm (micrometer) 1 µm = 1000 nm (nanometers) 1 nm = 1000 Å (angstrom) The human eye can see objects larger than 200–250 μm.  Light Microscope A standard microscopes have 4 objectives. Each objective has the magnification power engraved on the side: They will also be color coded for your convenience. Ø 4x: Scanning objective Ø 10x: Low power objective Ø 40x: High power objective Ø 100x: Oil immersion objective The Properites of Light Microscope Objectives Parts of Light Microscope Eyepiece (Ocular Lens) Diopter Adjustment Head Objective Lenses Nose Piece Arm(Frame) Stage Clips Stage Condensor(Diaphragm) Stage Control Light Switch Illuminator Fine Adjustment Knob (Light Source) Base Coarse Adjustment Knob Types of Microscope LIGHT MICROSCOPE : use sunlight or artificial light. 1. Bright field microscope. 2. Dark field microscope. 3. Phase contrast microscope. 4. Fluorescence microscope. ELECTRON MICROSCOPE : use of electron. 1. Transmission electron microscope. 2. Scanning electron microscope.  Choosing a microscope ØLook at organisms, cells or tissues that are currently alive? ØLook at the surface of a living thing? ØLook at whole cells and how they connect in 3D? ØLook at the surface of a sample at high resolution? ØLook at a cross-section of a sample at high resolution? ØBuild up a 3D model from the results? ØAvoid removing moisture from the sample? Examination of living cells Cell and Tissue Cultures  ATCC (American Type Culture Collection) Cell culture is the process of growing and maintaining cells outside their natural environment, typically in a laboratory setting. Cell culture Cells can be isolated from tissues, organs, or cell lines They cultivated in a controlled environment that provides the necessary nutrients, temperature, humidity, and other conditions for their growth and proliferation. Isolating cells and growing them in culture (Cell Culture) Living cells → can be suspended in an appropiate liquid Examined under the light microscope ↓ They will soon die Prolonged study of living cells can be made ↓ by culturing them in solutions (containing necessary nutrients) to keep them alive Cell Culture Conditions A medium that supplies the essential nutrients (amino acids, carbohydrates, vitamins, minerals Growth factors Hormones Gases (O2, CO2) A regulated physico-chemical environment (pH, osmotic pressure, temperature) Most cells are anchorage-dependent and must be cultured while attached to a solid or semi-solid substrate Mammalian cell culture medium Cell culture involves several key components: Rich media are required (contain necessary nutrients) Amino acids 10 amino acids called essential amino asids can not be synthesized by vertebrates and must be obtained from diet; Arginine, histidine, isoleucine, leucine, lysine, methionine, phenylalanine, threonine, tryptophan, valine) must be supplied Vitamins Various Salts Glucose Serum (contains various factors needed for proliferation of cells; growth factors) Mammalian cell culture medium Most animal cells can only grow on special solid surfaces (adhere to and grow on glass and treated plastics) Medium should be changed frequently Aseptic technique is necessary to avoid contamination (Antibiotics) We use a cell culture hood (laminar-flow hood) Temperature (37 ºC) pH 7,4 (phenol red: indicator dye) Primary cell culture Cell lines ØDerived from living tissue ØEukaryotic cell lines are a widely or an organism's organs used cell source for experiments ØCells isolated directly from ØAn immortalized or continuous human or animal tissue using cell line has acquired the enzymatic or mechanical ability to proliferate methods. indefinitely, either through genetic mutations or artificial ØLimited life span modifications. ØDivide ~ 25-50 times ØGrow indefinitely in culture ØStop dividing ØA culture derived from single transformed cell is called “ cell line” Applications of the cell culture Basic and Applied Aspects of Biotechnology pp 59–75  STEM CELLS A stem cell is a cell with the unique ability to develop into specialised cell types in the body. Toti = Whole; Pluri = Many; Multi = Several Journal of Clinical Medicine 8(5):627 Stem Cell Types Nat Rev Mol Cell Biol 22, 671–690 (2021)  Embriyonic stem cells Embryonic stem cells (ESCs) are cells derived from the inner cell mass of the blastocyst prior to implantation. They are pluripotent and have an unlimited capacity for self-renewal and the ability to differentiate into any somatic cell type. ES cells can be derived from early human embryos these cells might be used to replace and repair damaged mature human tissues. Induced Pluripotent Stem Cell: iPCS Hybrid cells Hybrid cells, also known as hybridomas in a specific context, have various applications in biomedical research, biotechnology, and medicine. The use of hybrid cells What is cell-cell fusion? Cell fusion: membrane merging and cytoplasmic mixing of two cell types 2 Types of Cell Fusion -Homotypic cell fusion: occurs between cells of the same type. -Heterotypic cell fusion: occurs between cells of the different type. Is cell fusion a normal process? We need specific substances that promote cell to cell fusion Hybridoma cell lines are factories that produce monoclonal antibodies B lymphocytes produce antibodies Problem; B lymphocytes have limited life span in culture To overcome this problem B lymphocytes from an immunized mouse are fused with cells derived from an immortal B lymphocyte tumor Hybrid cells have both the ability to make a specific antibody and to divide indefinitely (immortal) These hybridomas provide a permanent and stable source of a single type of monoclonal antibody Hybrid cell applications Hybridomas are primarily used for producing monoclonal antibodies, which are highly specific antibodies that can target a single antigen. 1) Disease diagnosis: to detect specific proteins or pathogens in patient samples.(HIV test, pregnancy test.. Etc.) 2) Therapeutics: cancer, autoimmune disorders, and infectious diseases. 3) Research 4) Vaccine Development:Some vaccines are produced using hybrid cell lines. 5) Immunotherapy:Chimeric antigen receptor (CAR) T-cell therapy Applications of cell fusion ü Nuclear transfer ü Embryo manipulation ü Hybridoma production Which cells are useful for this method ? Embriyonic cells Somatic cells Mammalian cell lines Oocytes and gametes Stem cells Cancer cells In vitro fertilization In vitro fertilization (IVF) is a process by which egg cells are fertilized by sperm outs