Unit 1: Cells PDF

Learning biology by numbers: Size and Geometry of cells, viruses and molecules; measurements in biology; Cellular building blocks: four classes of macromolecules, nucleic acids and proteins are polymer languages with different alphabets. Making a Cell: Construction of Cells and Organism - Spatial o...

Learning biology by numbers: Size and Geometry of cells, viruses and molecules; measurements in biology; Cellular building blocks: four classes of macromolecules, nucleic acids and proteins are polymer languages with different alphabets. Making a Cell: Construction of Cells and Organism - Spatial organization, Temporal organization.  Cell : The Unit of Life The smallest functional unit of life is cell, discovered by Robert Hooke in 1665. A cell can independently perform all necessary activities to sustain life. Hence cell is the basic unit of life. There are two types of cells → plant cell and animal cell. Cells have different organelles, each one with a distinct function. Size of cells vary greatly Generally small and seen only with microscope Relative sizes of the cells 3  Size and Geometry of cells The standard cells. (A) A schematic bacterium revealing the characteristic size and components of E. coli. (B) A budding yeast cell showing its characteristic size, its organelles, and various classes of molecules present within it. (C) An adherent human cell. (adapted from Alberts B, Johnson A, Lewis J et al. Molecular Biology of the Cell, 6th ed. Garland Science.) Both plant and animal cells show diverse shapes such as – Cell Organelles/Compartments - Plasma membrane – Boundary, Protection and transport Nucleus - hereditary data essential for multiplication and cell development. Endoplasmic Reticulum & Golgi apparatus- Protein processing and Lipid biosynthesis Mitochondria - Energy factories of cells Lysosomes - digest undesirable materials present in the cell Chloroplast (only Plant cells) – Photosynthesis Ribosomes – Protein synthesis Cell Theory Cells were discovered in 1665 by Robert Hooke. Early studies of cells were conducted by - Mathias Schleiden (1838) - Theodor Schwann (1839) Schleiden and Schwann proposed the Cell Theory. 9 Cell Theory Cell Theory 1. All organisms are composed of cells. 2. Cells are the smallest living things. 3. Cells arise only from pre-existing cells. All cells today represent a continuous line of descent from the first living cells. 10 Cell Theory Microscopes are required to visualize cells. Cell size is limited. -As cell size increases, it takes longer for material to diffuse from the cell membrane to the interior of the cell. Light microscopes can resolve structures that are 200nm apart. Electron microscopes can resolve structures that are 0.2nm apart. 11 Why are cells so small? Cells need to produce chemical energy (via metabolism) to survive and this requires the exchange of materials with the environment The rate of metabolism of a cell is a function of its mass / volume (larger cells need more energy to sustain essential functions) The rate of material exchange is a function of its surface area (large membrane surface equates to more material movement) As a cell grows, volume (units3) increases faster than surface area (units2), leading to a decreased SA:Vol ratio If metabolic rate exceeds the rate of exchange of vital materials and wastes (low SA:Vol ratio), the cell will eventually die Hence growing cells tend to divide and remain small in order to maintain a high SA:Vol ratio suitable for survival Cell Theory All cells have certain structures in common. 1. genetic material – in a nucleoid or nucleus 2. cytoplasm – a semifluid matrix 3. plasma membrane – a phospholipid bilayer 13 Cell Theory 14 Prokaryotic Cells Prokaryotic cells lack a membrane-bound nucleus. -genetic material is present in the nucleoid Two types of prokaryotes: -archaea -bacteria 15 Prokaryotic Cells Prokaryotic cells possess -genetic material in the nucleoid -cytoplasm -plasma membrane -cell wall -ribosomes -no membrane-bound organelles 16 Prokaryotic Cells Flagella -present in some prokaryotic cells -used for locomotion -rotary motion propels the cell 17 Eukaryotic Cells Eukaryotic cells -possess a membrane-bound nucleus -are more complex than prokaryotic cells -compartmentalize many cellular functions within organelles and the endomembrane system -possess a cytoskeleton for support and to maintain cellular structure 18 Eukaryotic Cells 19 Eukaryotic Cells 20 Eukaryotic Cells Nucleus -stores the genetic material of the cell in the form of multiple, linear chromosomes -surrounded by a nuclear envelope composed of 2 phospholipid bilayers -in chromosomes – DNA is organized with proteins to form chromatin 21 Eukaryotic Cells 22 Eukaryotic Cells Ribosomes -the site of protein synthesis in the cell -composed of ribosomal RNA and proteins -found within the cytosol of the cytoplasm and attached to internal membranes 23 Endomembrane System Endomembrane system -a series of membranes throughout the cytoplasm -divides cell into compartments where different cellular functions occur 1. endoplasmic reticulum 2. Golgi apparatus 3. lysosomes 24 Endomembrane System Rough endoplasmic reticulum (RER) -membranes that create a network of channels throughout the cytoplasm -attachment of ribosomes to the membrane gives a rough appearance -synthesis of proteins to be secreted, sent to lysosomes or plasma membrane 25 Endomembrane System Smooth endoplasmic reticulum (SER) -relatively few ribosomes attached -functions: -synthesis of membrane lipids -calcium storage -detoxification of foreign substances 26 Endomembrane System 27 Endomembrane System Golgi apparatus -flattened stacks of interconnected membranes -packaging and distribution of materials to different parts of the cell -synthesis of cell wall components 28 29 Endomembrane System Lysosomes -membrane bound vesicles containing digestive enzymes to break down macromolecules -destroy cells or foreign matter that the cell has engulfed by phagocytosis 30 31 Endomembrane System Microbodies -membrane bound vesicles -contain enzymes -not part of the endomembrane system -glyoxysomes in plants contain enzymes for converting fats to carbohydrates -peroxisomes contain oxidative enzymes and catalase 32 Endomembrane System Vacuoles -membrane-bound structures with various functions depending on the cell type There are different types of vacuoles: -central vacuole in plant cells -contractile vacuole of some protists -vacuoles for storage 33 Mitochondria Mitochondria -organelles present in all types of eukaryotic cells -contain oxidative metabolism enzymes for transferring the energy within macromolecules to ATP -found in all types of eukaryotic cells -surrounded by 2 membranes -smooth outer membrane -folded inner membrane with layers called cristae -matrix is within the inner membrane -intermembrane space is located between the two membranes -contain their own DNA 34 Mitochondria 35  Chloroplasts of Mitochondria of Prokaryotes Eukaryotes Photosynthetic Eukaryotic cells eukaryotes Multiple linear 1 single, circular chromosomes 1 single, circular 1 single, circular DNA chromosome compartmentalized in chromosome chromosome a nucleus Binary Fission Binary Fission Binary Fission Replication Mitosis Ribosomes "70 S" "80 S" "70 S" "70 S" Found in the plasma Not found in the Found in the plasma Found in the Electron Transport membrane around plasma membrane membrane around plasma membrane Chain cell around cell mitochondrion around chloroplast Size (approximate) ~1-10 microns ~50 - 500 microns ~1-10 microns ~1-10 microns Anaerobic bacteria: ~3.8 Billion years Appearance on Photosynt.bacteria: ~1.5 billion years ~1.5 billion years ~1.5 billion years ago Earth ~3.2 Billion years ago ago Aerobic bacteria: ~2.5 Billion years 36 Chloroplasts Chloroplasts -organelles present in cells of plants and some other eukaryotes -contain chlorophyll for photosynthesis -surrounded by 2 membranes -thylakoids are membranous sacs within the inner membrane -grana are stacks of thylakoids 37 Chloroplasts 38 Mitochondria & Chloroplasts Endosymbiosis -proposal that eukaryotic organelles evolved through a symbiotic relationship -one cell engulfed a second cell and a symbiotic relationship developed -mitochondria and chloroplasts are thought to have evolved this way 39 Mitochondria & Chloroplasts Much evidence supports this endosymbiosis theory. Mitochondria and chloroplasts: -have 2 membranes -possess DNA and ribosomes -are about the size of a prokaryotic cell -divide by a process similar to bacteria 40 Cytoskeleton Cytoskeleton -network of protein fibers found in all eukaryotic cells -supports the shape of the cell -keeps organelles in fixed locations -helps move materials within the cell 41 Cytoskeleton Cytoskeleton fibers include -actin filaments – responsible for cellular contractions, crawling, “pinching” -microtubules – provide organization to the cell and move materials within the cell -intermediate filaments – provide structural stability 42 Cytoskeleton 43 Cell Movement Cell movement takes different forms. -Crawling is accomplished via actin filaments and the protein myosin. -Flagella undulate to move a cell. -Cilia can be arranged in rows on the surface of a eukaryotic cell to propel a cell forward. 44 Cell Movement The cilia and flagella of eukaryotic cells have a similar structure: 9+2 structure: 9 pairs of microtubules surrounded by a 2 central microtubules Cilia are usually more numerous than flagella on a cell. 45 Cell Movement 46 Extracellular Structures Extracellular matrix (ECM) -surrounds animal cells -composed of glycoproteins and fibrous proteins such as collagen -may be connected to the cytoplasm via integrin proteins present in the plasma membrane 47 Extracellular Structures 48 49  How cells are constructed? ▪ New cells are created from existing cells through a process referred to as the cell cycle. One cell can make a copy of itself and form two new daughter cells. ▪ There are two major tasks that have to happen every cell cycle. First, cells have to make an exact copy of their DNA. DNA is like the instruction manual for a cell. It encodes genes for characteristics and dictates things like eye color and blood type. ▪ The second major task of every cell cycle is for the replicated chromosomes to be organized and separated into opposite sides of the cell. This happens during mitosis, or M phase of the cell cycle. ▪ The cell then grows longer, further separating those masses of chromosomes. The middle of the cell then pinches off in a process known as cytokinesis, splitting the cell into two cells. A new cell has been created and that completes the cell cycle.  How cells build Organisms ? Organizational control mechanism allows cells to form tissues and anatomical structures in the developing embryo Spatial and Temporal organization ▪ Individual-specific functional brain networks can be elucidated in the cerebellum ▪ Precision functional mapping of individuals revealed that functional networks in the cerebral cortex exhibit measurable individual specificity. Cell Census An order-of-magnitude census of the major components of the three model cells Importance of cell census Realistic physical picture of any biological phenomenon demands a precise, quantitative understanding of the individual molecules involved and the distance between them You will find the cell interior is extremely crowded in contrast to the dilute and homogeneous environment of the biochemical test tube. We will see that the mean spacing between protein molecules within a typical cell is less than 10 nm. This is extremely useful to estimate the rates of macromolecular synthesis during the cell cycle. E Coli: a model organism which has led to astounding discoveries Is easy to isolate present in human fecal matter E. coli is able to grow well in the presence of oxygen Easy to culture in lab, has high growth rate Genome has been sequenced 1997 It carries plasmids extra-chromosomal DNA which can be manipulated easily by molecular biology techniques. Molecular biology techniques are easy to apply for creation of mutants Often, we will have recourse to E. coli because of particular experiments that have been performed on this organism. Further, even when we speak of experiments on other cells or organisms, often E. coli will be behind the scenes coloring our thinking.  Size of an E. coli cells and molecular composition E. coli. are made up of an array of different macromolecules as well as small molecules and ions. To estimate the number of proteins in an E. coli cell We begin by noting that with its 1 fL volume, the mass of such a cell is roughly 1 pg, where we have assumed that the density of the cell is that of water which is 1 g/mL. Measurements reveal that the dry weight of the Cell is roughly 30 percent of its total and half of that mass is protein. As a result, the total protein mass within the cell is roughly 0.15 pg We can also estimate the number of carbon atoms in a bacterium on the grounds that roughly half the dry mass comes from the carbon content of these cells, a figure that implies 1010 carbon atoms per cell Revealing the extent of crowding within a bacterium, we can estimate the number of proteins by assuming a mean protein of 300 amino acids with each amino acid having a characteristic mass of 100 Da. Using these rules of thumb, we find that the mean protein has a mass of 30,000 Da. Using the conversion factor that 1 Da 1.6 × 10−24 g, we have that our typical protein has a mass of 5 × 10−20 g. The number of proteins per E. coli cell  How big is an E. coli cell ? ▪ The size of a typical bacterium such as E. coli serves as a convenient standard ruler for characterizing length scales in molecular and cell biology ▪ Diameter ≈1μm, a length of ≈2μm, and a volume of ≈1μm3 ▪ The shape can be approximated as a spherocylinder—that is, a cylinder with hemispherical caps. ▪ Inferences can vary with cell types under various conditions. Relation between cell volume and growth rate. Using microscopy and microfluidic devices, cell volume can be measured at the single-cell level under various conditions, confirming that the average cell volume grows exponentially with growth rate.  =10-3Pa⋅sec =147bp 1 Da = (mass of one carbon-12)/12  E V ≈ 1μm3 = 1 fL ≈ 1pg.Coli m E.Coli ≈ 1g / mL ≈ ρwater ρ E.Coli 1mol molecule / 1 fL 6 × 1023 × 10−15 ≈ 2nM 1 = L Thermal energy scale (at 300K) k B T = 4.14 × 10−21J = 4.14 pN ⋅ nm AB  A + B  0.59kcal / mol [A][B] ΔG = k B T log d = k BT log ≈ k BT log10 −9= − 2.3 × 9kB T ≈ 20k BT K [AB] − ≈ 20kBT (Kd = 1nM )[1molecule / Typical protein-protein cell] ≈ 14kB T (K 3 d = 1μM )[10 molecules / cell] interaction energy ≈ 7kBT (Kd = 1mM )[106 molecules / cell]  (m Water is 70 % of the cell mass E.Coli = 1pg) Dry mass of the cell (30% of 1pg) = 0.3 pg Half of the dry mass (=0.15 pg) = proteins 1 g = 1.6 × 10−24 g 1 Da = mass of a hydrogen atom = 6 × 1023 1 amino acid = 100 Da average protein size = 300 a.a. → 30,000 Da 0.15 0.15 pg Nprot= = ≈ 3, 000, pg 30, −24 30, 000 × 1.6 × 10 000 ⎧ 1N → 000Da g membrane proteins mwater= 0.7 pg ⎪ prot → ⎨ 3 ⎪ 2 N prot → cytoplasmic proteins 0.7 N water= ≈2× ⎪ pg× 10−24 g / 18Da × (1.6 ⎩ 1010 Da)  6 There are proteins in cytoplasm 2 × 10 c = 2 × 106 / 1μm3 prot ⎛ ⎞ 103 nm 3⎥ 1/ 3 = 500nm d prot − prot =c −1/ 3 = ⎜( ) ( )1/ 3 ≈ 8nm ⎜ 2 × 106 ⎝ 4 nm 4 nm ⎥ ⎠ 8 nm Mean spacing between the macromolecules in cell ~ size of macromolecules themselves Cell is very crowded... 28 Membrane Surface area of E.Coli A ≈ (2πR) × L ≈ 6μm2 E.Coli E. coli has double (inner and outer) membranes and each membrane is made of bilayer. Half of the surface area is covered with membrane proteins 4 × 0.5 × 6μm2 lipid E ≈ ≈ 2 × 10 7 N.Coli 0.5nm2 29 Taking the molecular census to set criteria for the judgement Measurement of protein census of E. Coli using 2D polyacrylamide gel electrophoresis Experimental determination of biomolecular content Large and negatively charged proteins 1.Protein mixture loaded at one end of the gel and an electric field is applied across the gel. (F=qE=ζv) 2.A charged detergent binding to proteins is added. (charge of detergent ~ protein size; q ~ R~v) 3. Proteins are stained using dye. Conc ~ Intensity of the spot. 4.Cut each spot, elute the proteins and determine the size and amino acid content using mass spectroscopy. small and positively charged proteins RIBOSOME 20% of the protein complement of a cell = ribosomal proteins Ribosome (70S) = Large subunit + small subunit Large subunit (50S) = 23S r-RNA + r-proteins Small subunit (30S) = 16S r-RNA + r-proteins S : Svedberg constant (sedimentation constant) - A heavier particle sediments faster in the centrifugation, thus have a larger S value. m(1 − ρν )rω = ξ dr 2 dt M (1 − ρν )D d log r = ≡ RT ω 2 dt S ⎧ r − RNA : 2/3 of the mass ribosome (2.5MDa) ⎪ ⎨⎪ r − protein : 1/3 of the mass ⎩ M ∴ N ribosome r − prot = 0.2 × 0.15 pg = 20, 000 mr − protein = 830, 000Da = m r− 830, 000Da M r − protein = 20% × M ≈ 19, prot protein Conversion : Number ➯ Concentration ➯ Average distance * 1 molecule in a bacterium ~ 2 nM. Example : Ribosome 1L = 10−3 m3 no. of ribosome = 19,000 (Table 2.1) Concentration = 19,000/1fL ~ 32 μM Average distance ~ 37 nm. Cellular crowding and its implications The Cellular Interior Is Highly Crowded With Mean Spacings Between Molecules That Are Comparable to Molecular Dimensions Increase in the effective concentration of macromolecules alters the rates and equilibrium constants of their reactions Alters dissociation constants by favoring the association of macromolecules, such as when multiple proteins come together to form protein complexes, or when DNA-binding proteins bind to their targets in the genome Crowding may also affect enzyme reactions involving small molecules if the reaction involves a large change in the shape of the enzyme. The size of the crowding effect depends on both the molecular mass and shape of the molecule involved,  Cellular crowding and its implications the increase in the strength of interactions between proteins and DNA is importance in processes such as transcription and DNA replication involved in processes as diverse as the aggregation of hemoglobin in sickle-cell disease, and the responses of cells to changes in their volume. the crowding effect can accelerate the folding process, crowding can reduce the yield of correctly folded protein by increasing protein aggregation.[ increase the effectiveness of chaperone proteins such as GroEL in the cell, Crystallins fill the interior of the lens. These proteins have to remain stable and in solution for the lens to be transparent; precipitation or aggregation of crystallins causes cataracts Crystallins are present in the lens at extremely high concentrations, over 500 mg/ml, and at these levels crowding effects are very strong. The large crowding effect adds to the thermal stability of the crystallins, increasing their resistance to denaturation This effect may partly explain the extraordinary resistance shown by the lens to damage caused by high temperatures. Crowding may also play a role in diseases that involve protein aggregation, such as sickle cell anemia alzheimer's disease, Biological Structures exist over a huge range of scales Hierarchy of spatial scales Number of proteins in an E.coli cell 3,000,000 Number of ribosomes in an E.coli cell 20,000 Number of lipids in an E.coli cell 20,000,000 Size of genome in an E.coli cell 5,000,000 bp  yeast cell : useful representative to study eukaryotes Tremendous diversity in living cells... Protist Cells Biological Structures exist over a huge range of scales Hierarchy of spatial scales yeast cell : model system to study a single eukaryote cell Lyeast 5μm V = 4 π 2.5μm 3 ≈ 3 ≈ 60VE yeast 3 ( ).Coli ≈ 60 × N60 μm E.Coli Yeast protein N protein Yeast ≈ 2 × 0.5 × 80 μm2 ≈ 2 × 108 N lipid 0.5nm2 ( 1.2 × 107 bp ) 7 N genome ~ 1.2 × 10 bp N nucleosome  200bp / = 60, 000 nucleosome  Video resources https://www.youtube.com/watch?v=URUJD5NEXC8 https://dnalc.cshl.edu/resources/3d/08-how-dna-is-packaged-advanced.html https://www.youtube.com/watch?v=jOhNyVjkChM 43 VIRUSES A virus is an infectious microbe consisting of a segment of nucleic acid (either DNA or RNA) surrounded by a protein coat. A virus cannot replicate alone; instead, it must infect cells and use components of the host cell to make copies of itself. Often, a virus ends up killing the host cell in the process, causing damage to the host organism. Well-known examples of viruses causing human disease include AIDS, COVID-19, measles and smallpox. ▪ Viruses are much smaller than the cells they infect. ▪ They could pass through filters small enough to remove pathogenic bacterial cells. ▪ These genomes can be DNA or RNA, single-stranded or double-stranded (that is, ssDNA, dsDNA, ssRNA, or dsRNA) with characteristic sizes ranging from 103–106 bases (A) Electron microscopy image of phi29 and T7 bacteriophages as revealed by electron microscopy. (B) Schematic of the structure of a bacteriophage. (A, adapted from Grimes S, Jardine PJ & Anderson D Adv Virus Res 58:255–280.) Cellular building blocks ▪ Cells of all organisms consist of four fundamental macromolecular components: nucleic acids (including DNA and RNA), proteins, lipids and glycans. ▪ From the construction, modification and interaction of these components, the cell develops and functions. ▪ DNA and RNA are produced from the 8 nucleosides. Although deoxyribose (d) and ribose (r) are saccharides, they are an integral part of the energetically charged nucleoside building blocks that are used to synthesize DNA and RNA. ▪ There are 20 natural amino acids used in the synthesis of proteins. ▪ Glycans derive initially from 32, and possibly more, saccharides used in the enzymatic process of glycosylation and are often attached to proteins and lipids. ▪ Lipids are represented by 8 recently classified categories and contain a large repertoire of hydrophobic and amphipathic molecules. The central dogma of Life