Lesson-2-CELL.pdf

BIOCHEMISTRY Lesson 2  THE CELL ❖All organisms are made up of cells. “ It is the basic structural, functional and biological unit of life.” ❖It was discovered by Robert Hooke in 1665. ❖The word cell comes from a Latin word 'cellula' meaning small room. ❖Cells are t...

BIOCHEMISTRY Lesson 2  THE CELL ❖All organisms are made up of cells. “ It is the basic structural, functional and biological unit of life.” ❖It was discovered by Robert Hooke in 1665. ❖The word cell comes from a Latin word 'cellula' meaning small room. ❖Cells are the building block of life, where all the chemical processes occur. ❖They take up the biological molecules, convert them into energy or use them to carry out specialized functions. ❖It also contains the hereditary material which determine the genetic characteristics. ❖The cell theory was developed by Matthias Jacob Schleiden and Theodor Schwann. It states that.. “All organisms are composed of one or more cells, and that all cells come from pre-existing cells. The vital functions of an organism occur within the cells and that all cells contain hereditary information necessary for cell functions and for transmitting from one generation to the next.” ❖Every living things (animals, plants, bacteria, fungi, protozoans) are made up of cell. Some organisms are made up of just one cell. ( Unicellular organism. e.g. Bacteria) While some organisms are made up of many cells. (Multicellular organisms e.g. Animal cell) ❖The human body is built with about 30 to 40 trillion cells specializing in different functions. ❖Many identical cells joins together and forms a tissue. ❖Various tissues, that perform a particular function, organized together which forms the organ. ❖Various organs joined together forms an organ system. TYPES OF CELL 1. Eukaryotic cell 2. Prokaryotic cell  1. EUKARYOTIC CELL Eukaryotic cells are those cells which have a true nucleus. It has a nuclear membrane within which there is well defined chromosomes. It has other membrane bound organelles like mitochondria, endoplasmic reticulum, Golgi bodies etc.  Organisms with eukaryotic cells are called as Eukaryotes. They may be single-celled or multicellular organisms. e.g. cells of plant, animals, fungi.  2. PROKARYOTIC CELL Prokaryotic cells are those cells whose nucleus is not distinct and their DNA is not organized into chromosomes. They lack most of the membrane bound organelles. Prokaryotic cells are also called Prokaryotes. They are usually unicellular organisms. e.g. bacteria.  EUKARYOTIC CELL A typical Eukaryotic cell, as seen under light microscope has two major parts: The cell membrane The cytoplasm and its organelles The different substances that make up the cell are collectively called as Protoplasm.  1. CELL MEMBRANE It is a thin, elastic, semi permeable membrane of 7.5 to 10 nanometers thickness. It is a living membrane made up of phospholipid bilayer embedded with proteins. It is a flexible membrane and so it can fold in or out. It is made up of 55 % proteins, 25 % phospholipids, 13 % cholesterol, 4% lipids and 3% carbohydrates. Functions of the cell membrane It protects the cell contents from the surrounding environmement. It provides a shape to the cell. It is semi permeable and allows transport of certain substances into and out of the cell. It helps in forming cell junctions. 2. CYTOPLASM AND ITS ORGANELLES In eukaryotes, the protoplasm surrounding the nucleus is called cytoplasm. It is a clear gelatinous fluid that fills the cell and surrounds the organelle. It contains 90 % water, dissolved substances, minerals, sugar, irons, vitamins, amino acids, proteins and enzymes.  Cytoplasm is the seat for many pathways like glycolysis and HMP (Hexose monophosphate) pathway. The cytoplasm contains well organized structures called organelles which vary in size from a few nanometers to many micrometers and they are specialized to carry out one or more vital functions of the cell. Cytoplasm vs cytosol?  The organelles include... –Mitochondria –Endoplasmic reticulum –Golgi apparatus –Ribosomes –Lysosomes –Peroxisomes –Centriole –Vacuole –Nucleus  These are tiny, sausage shaped structures of diameter 0.5 to 1 micrometer. It is called as “Power house of the cell” as it generates energy in the form of AdenosineTriphosphate (ATP) which is required by all the cells. It is found both in plant and animal cells. It has a double membrane envelope: an inner membrane and an outer membrane.  The inner membrane is folded and pleated (double fold) and it is called cristae. It provides large surface area for different biochemical processes as it contains many oxidative enzymes. Within this membrane are the proteins involved in electron transport chain, ATP synthase and transport proteins. It is impermeable to molecules and ions but allows the free passage of carbon dioxide, oxygen and water.  The metabolites are transported across the membrane with the help of transport proteins. The outer membrane is a smooth phospholipid bilayer enveloping the mitochondria. It had enzymes like monoamine oxidase and NADH reductase. (NADH- Nicotinamide adenine dinucleotide + Hydrogen) An intrinsic protein called porin form the channels that makes the membrane permeable to solutes and metabolites.  It allows the free passage of substances with molecular weight of less than 10,000. The membranes create two compartments. –The space between the outer and inner membrane is called the intermembrane space. –It is here that oxidative phosphorylation occus. (Releasing energy by oxidizing nutrients for ATP synthesis) –The inner cavity of mitochondria is called the matrix.  It is packed with many enzymes like pyruvate dehydrogenease, pyruvate carboxylase, enzymes for oxidation of fatty acids, aminoacids and enzymes of citric acid cycle. It also contains the mitochondrial genome, mitochondrial ribosomes, tRNAs, dissolved oxygen, carbon dioxide and water.  FUNCTIONS:- –It is the seat for the Kreb's cycle (citric acid cycle) –It contains enzymes for Oxidative phosphorylation which helps in producing energy rich ATP molecules –It provides intermediates for synthesis of cytochrome, chlorophyll, hemoglobin and steroids –Amino acids like glutamate are synthesized in it from alpha ketoglutarate and oxalo acetate –Many fatty acids are synthesized in the matrix –Calcium can be stored in the mitochondria and released whenever required.  It is a network of tubular and vascular structures extending from outer membranes of nucleus to the cell membrane. It is seen to be spread throughout the cytoplasm and it provides a large surface area for various physiological activities. The inside of the vesicles and tubules is filled with an endoplasmic matrix.  When the ribosomes are attached on the outer surface of mebrane of the endoplasmic reticulum, it is called as Rough endoplasmic reticulum (RER). It lies adjacent to the cell nucleus and its membrane is continuous with the membrane of the nucleus. When there are no ribosomes attached to the endoplasmic reticulum, it is called as Smooth endoplasmic reticulum (SER).  Functions:- –RER helps in transporting proteins from ribosomes to Golgi bodies. –Proteins that enters RER undergoes processing, folding and sorting –SER is involved in the synthesis of lipids, including cholesterol and phospholipids –In some cells, SER helps in the synthesis of steroid hormones from cholesterol –In the cells of liver, SER helps in detoxifying drugs and harmful chemicals.  It is a stack of membranous sac, like a pile of discs. It is present between endoplasmic reticulum and cell membrane. Like endoplasmic reticulum, it is a single membrane bound structure. In animal cells, it is present around the nucleus while in plant cell it is scattered throughout the cell. These cell organelles pack and sort the proteins before they are sent to their destinations.  Functions:- –The proteins that enter it from RER is modified, processed, sorted and transported in the form of vesicles to the cell membrane and other destinations. –Glycolipids, sphingomyelin are synthesised within it. –In plant cells, it help in the synthesis of polysaccharides needed by the cell wall. –It has a role in the synthesis of carbohydrates like galactose. –Primary lysosomes develop from mature Golgi bodies. –It plays an important role in lipid trafficking.  They are spherical shaped organelles seen either free in the cytoplasm or attached to RER. They are found in eukaryotes and prokaryotes. They are synthesised by the nucleolus. The ribosomes link the amino acids together in the order that is specified by the messengers RNA. They are made up of two subunits - a small sub unit and a large sub unit.  The small sub unit reads the mRNA while the large subunit assembles the amino acids to form large polypeptide. The ribosomal sub units are made up of one or more eRNA and proteins. Function:- –It is the site for protein synthesis.  They are tiny sac like organelles of size 0.5 to 1.5 µm, which are membrane bound and found in hundreds in a single cell. They are formed from the Golgi bodies as small vesicles which bud off from them. Within the sac there are several hydrolytic enzymes that breakdown macromolecules like nucleic acid, proteins and polysaccharides.  They are also called as “suicidal bags” as enzymes contained in them can digest the cell's own material when damaged or dead. The important enzymes in it are DNA- ase, RNA-ase, protease, lipase, glycosidase, phosphatase, sulfatase which are synthesized in the endoplasmic reticulum and then transported to the Golgi bodies.  Functions:- –It helps in the digestion of food releasing enzymes. –They digest worn out organelles –It helps in the defense, by digesting germs –It helps sperm cells in entering the egg by breaking through the egg membrane –It provides energy during cell starvation by the digestion of its own cell parts.  It is a type of microbody which is small, spherical shaped, with single membrane and of size 0.5-1.5 µm. They are found in both animal and plant cells. Just like lysosomes they contain many enzymes which help in biological reactions. They are formed from the endoplasmic reticulum unlike lysosomes which are formed from the Golgi bodies.  Functions:- –The enzymes found in peroxisomes are usually used for different metabolic reactions and for digesting different materials in the cell. –They help in the oxidation of many substances resulting in the formation of hydrogen peroxide as a by product. –But, it contains enzyme peroxidase or catalase which decomposes this harmful hydrogen peroxide into water and oxygen or uses it to oxidize other organic compounds like phenol, alcohol, formaldehyde, etc. –They are also involved in the catabolism of fatty acids (beta oxidation), D- aminoacids and polyamines. –They are needed in the synthesis of plasmalogens (type of ether phospholipid needed for functioning of brain and lungs.) –They participate in the synthesis of cholesterol, bile acids and myelin. –In plants, it helps in the photorespiration and symbiotic nitrogen fixation.  They are long, hollow cylinders of size 24 nm in diameter and can grow up to a length of 50 µm. They are found in eukaryotic cells.  They are made up of two globular proteins namely α-tubulin and β-tubulin. Along the microtubule axis tubulins are joined end to end to form protofilaments.  The cytoskeleton is a structure that helps cells maintain their shape and internal organization, and it also provides mechanical support that enables cells to carry out essential functions like division and movement. They are organized by microtubule organizing structures, primarily the centrioles.  Functions:- –They are part of the cytoskeleton and it provides mechanical support to the cell –It helps in the organization of cytoplasm –They help in the segregation of chromosomes during mitosis –They are used for locomotion (movement from one place to another) when present)  These are small rod like structures of size 4-7nm in diameter found in the cytoplasm of all eukaryotic cells, forming a part of the cytoskeleton. They are made up of protein, Actin (contractile protein) Functions:- –It provides support and shape to cell –Along with myosin, it helps in contraction –It helps in cytokinesis (a physical process of cell division)  They are part of the cytoskeleton in the cytoplasm, also surrounding the nucleus and extending to the cell membrane. They are made up of different types of fibrous proteins unlike microtubules which are made up of actin. They are of size 8-12 nm in diameter. They are found in hair, nails, scales and skin since they have high tensile strength.  Due to their rope like structure they provide mechanical strength to the cells and help cells to withstand stress like stretching and changing shape. e.g. Keratin filaments in skin and epithelial cells.  It is cylindrical in shape and of length 0.5 micrometer. it is present in all animal cells just outside the nucleus. It does not have a membrane. All centrioles are made up of protein strands called tubulin. Each centriole has 9 sets of inter connected peripheral tubules and each set has 3 micro tubules arranged at definite angles making the shape of a cylinder.  It has its own DNA and RNA and therefore, self duplicating. Two centrioles when oriented at a right angle, forms the centrosome. ❖Functions:- ▪It is involved in cell division. They are seen in the process of both meiosis and mitosis. ▪ It helps in the formation of cilia and flagella. ▪It helps in the organization and alignment of microtubules within the cell.  It is a membrane enclosed fluid filled sac present in animal and plant cells including fungi. It contains organic and inorganic molecules within it. They do not have particular size or shape but adjust themselves according to the need of the cell. ❖Functions:- ▪It helps in removing waste products from the cell ▪It isolates substances that are harmful to the cell ▪ It holds water and waste products within it ▪ It helps in maintaining the internal pH of cell ▪It helps to maintain hydrostatic pressure within the cell ▪It plays a major role in autophagy by maintaining a balance between biogenesis and degradation. NUCLEUS  It is a specialized double membrane- bound protoplasmic body present at the center of the cell. It is known as the cell's information center as it houses the chromosomes. The double membrane around the nucleus is called nuclear membrane or nuclear envelope. It is made of proteins and lipids, It enclose the nucleus to keep it separate from surrounding materials off the cell.  The outer membrane is continuous with endoplasmic reticulum and it has ribosomes attached on the outer surface. It has several large nuclear pores through which nuclear transport of large molecules, small molecules and ions occur. The space between the nuclear membranes is called perinuclear space and it is continuous with the lumen of rough endoplamic reticulum.  Within the nuclear membrane is a jelly like substance called karyolymph or nucleoplasm. Within it, there is a network of chromatin fibrils which condense to form chromosomes during cell division The nucleolus is present within the nucleus. It does not have a membrane around it. It synthesizes rRNA and assembles it. It regulates the synthetic activity of nucleus. ❖Functions:- ▪It controls the hereditary charateristics of an organism ▪Protein synthesis, cell division, growth and dfferentiation occurs in it ▪ Stores heredity materials in the form of DNA ▪It is the site for transcription in protein synthesis ▪Nucleolus helps in the synthesis of ribosomes ▪It regulates the integrity of genes and gene expression  A prokaryote is a single-celled organism that does not have a 'true nucleus'. e.g. Bacteria, Archaea. It does not contain any membrane bound organelles like mitochndria, nucleus, endoplasmic reticulum etc. They have many ribosomes scattered throughout their cytoplasm and nucleoid which contains the DNA.  Parts of Prokaryotic Cell: o Flagellum: –It is long whip like structure that helps in locomotion o Pili: –Small hair like structure present on the surface which helps in attaching to the surface of other bacteria. o Cell membrane: –It surrounds the cytoplasm and regulates the flow of substance in and out of the cell o Capsule: –It is a polysaccharide layer that is outside the cell envelope. –It enhances the ability of bacteria to cause disease o Cell wall: –It is the outer most covering of the cell and it gives shape to the cell o Cytoplasm: –It is gel like substance present within the cell. –It contains enzymes, ions, organic molecules, ribosomes, nucleoid o Ribosome: –It is the organelle which helps in protein synthesis. o Nucleoid: –It contains the genetic material o Plasmid: A small DNA molecule within the cell that can replicate itself. It is a small, double stranded and circular in shape organelle. These are usually found in bacteria.  MICROSCOPY It is the technical field of using microscopes to view samples and objects that cannot be seen with the unaided eye. It is the science of investing small objects using microscopes.  MICROSCOPE Microscope is an instrument used to see objects that are too small for the naked eye. Identification of minute organisms are necessary for diagnosis and treatment.  TYPES OF MICROSCOPE ❖Optical / Light microscope ❖Electron microscope ❖Darkfield or Ultra microscope ❖Phase Contrast and Differential Interference Contrast microscope ❖Fluorescent microscope ❖Ultraviolet microscope 1.OPTICAL/LIGHT MICROSCOPE ❖It a type of microscope which uses visible light and a system of lenses to magnify images of small samples. ❖It was invented by Hans and Zacharias Janssen in 1590. ❖The eyepiece, objective lenses, reflector, condenser and stage is first cleaned by a lens paper. ❖The specimen slide is then placed on the stage with help of clips at the center. ❖Observe through the eyepiece and focus the object with the help of coarse or fine adjustment. ❖The reflector can be adjusted to get proper light. ❖The light shining through the specimen is focused by the lens so that a magnified image can be seen through the eyepiece.  2. ELECTRON MICROSCOPE ❖It was designed by Ernst Ruska and Max Knoll in 1931 in Germany. ❖It is a type of microscope that uses a beam of electrons to illuminate a specimen and produces a magnified image. ❖The wavelength of lens as when compared to light is smaller and so they help magnify even very small objects. ❖It has high magnifying power when compared to light microscope as electrons have shorter wavelength than visible light. ❖It uses electrostatic and electro magnetic lenses to control electron beam and focuses in to form an image. ❖It is used to see microorganisms, cell organelles, large molecules, biopsy samples, etc.  TYPES OF ELECTRON MICROSCOPE ❖Transmission Electron microscope ❖Scanning Electron microscope ❖Reflection Electron microscope ❖Scanning Transmission Electron microsope ❖Low Voltage Electron microscope ❖Transmission Electron Microscope:- ▪It uses high voltage electron beam to create an image. ▪The beam of electron is sent through the specimen. ▪Some electrons are reflected while others pass through it creating an image of the specimen. ❖Scanning Electron Microscope:- ▪It does not produce a complete image of the specimen. ▪It scans the surface of the specimen and forms an image by detecting electron that are reflected or absorbed. ❖Reflection Electron Microscope:- ▪Similar to the TEM, the reflected electrons are detected to get the information about the surface of the specimen. ❖Scanning Transmission Electron Microscope:- ▪It combines high magnification of TEM with surface details of SEM. It helps to perform a complex analysis of the specimen. ❖Low Voltage Electron Microscope:- ▪It operates at accelerating voltage of a few kilo electrovolts or less.  3. DARKFIELD MICROSCOPE ❖A dark field microscope is arranged so that the light source is blocked off, causing light to scatter as it hits the specimen. ❖Here reflected light is used in place of transmitted light. ❖The oblique beams of refracted and diffracted light are coming from the sides and passes into and over the specimen to illuminate it. ❖The object looks bright on a dark background. ❖It is useful in observing small living objects and small organelles like nucleus, mitochodria, vacuole, etc. ❖It is usually used to see unstained objects. 4. PHASE CONTRAST MICROSCOPE ❖It is a microscope which helps to see unstained micro organisms. ❖It is the technique that converts phase shifts in light passing through a transparent specimen to brighness changes in the image. ❖The light passing through two different materials with different refractive index will undergo a change in the phase of light. ❖These phase differences are converted to difference in intensity of light, making image appear dark against a light background. ❖Phase contrast microscopy improves the contrast and make the structures visible.  5. FLUORESCENT MICROSCOPE ❖It is an optical microscope that uses fluorescence and phosphorescence to study the properties of organic and inorganic substances. ❖Special dyes like fluorescein, rhodamine and auramine are used.  6. ULTRAVIOLET MICROSCOPE ❖It is a microscope that has quartz lens and slides that uses ultra violet light as illumination instead of the common light. ❖Uses the shorter wavelength (180-400 nm) of ultraviolet rays compared to common light, which brings about higher resolution of objects.  CELL FRACTIONATION It is the technique of rupturing the cell to separate various cell components while preserving their individual functions in order to study their structure and chemistry. Cells can be broken down by many ways like subjecting it to osmotic shock or ultrasonic vibration or ground in a small blender. Albert Claude (1899-1983) pioneered techniques of cell fractionation by differential centrifugation.  These procedures break up the cell in to fragments and its nuclei, mitochondria, Golgi body, lysosomes, peroxysomes, etc. can be separated. The cells are first suspended in 0.25 molar sucrose solution at 0-4℃. Cells are then grounded well to form an isotonic slurry called homogenate. The homogenate is then subjected to different values of centrifugal force.  This helps in seperating the cell components by size and density. At a relatively low speed, the large components like nuclei sediments form a pellet at the bottom of the centrifuge tube, at slightly higher speed, a pellet of mitochondria is formed, and at even higher speeds and with longer period of centrifugation, first the small closed vesicles and then the ribosomes can be collected.  GEL ELECTROPHORESIS It is a method of separating macromolecules like DNA,RNA and proteins based on their size and charge, by passing it through a gel medium namely agarose and by applying an electric field. The smaller molecules move faster than the larger ones through the pores of the gel and the molecules in the gel can be stained to make them visible.  Ethidium bromide is the most commonly used stain to make DNA or RNA strands visible.  CHROMATOGRAPHY It is the technique used for separation of a mixture by dissolving it in a solution or suspension and allowing it to pass through a medium in which the components move at different rates. In this technique there is a stationary phase and mobile phase. The various constituents of the mixture travel at different speeds, causing them to separate.  TYPES OF CHROMATOGRAPHY ❖Column chromatography ❖Paper chromatography ❖Thin layer chromatography ❖Gel filtration chromatography ❖Ion exchange chromatography ❖Affinity chromatography