Week 2 Notes
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This 10 page Class Notes was uploaded by Madison Notetaker on Thursday February 4, 2016. The Class Notes belongs to BIO 310 at Missouri State University taught by Dr. Schweiger in Spring 2016. Since its upload, it has received 12 views. For similar materials see Microbiology in Biology at Missouri State University.
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Date Created: 02/04/16
1-20-16 Prokaryotic cell walls: Function: protection from osmotic pressure (pressure from inside and outside – turgor pressure) Major component is peptidoglycan- unique to bacteria made of glycan chain cross-linked by a short peptide Major component of the cell wall of archaea is pseudomurein (similar, murein is a component of peptidoglycan, but they are not the same) Bacterial Peptidoglycan Glycan chain made of two amino sugar derivatives - N-acetylglucosamine - N-acetylmuramic acid The glycan chain is connected by a short tetrapeptide that contains some unique amino acids (D-alanine, diaminopimelic acid, D-glutamate) ^^picture in notes Linkages between polysaccharides (backbone) and the tetrapeptides are always from muramic acid regardless of the bacteria. Lysozyme: attacks the cells wall NAG-NAM linkage (Alexander Fleming) B-lactams (Penicillin): inhibits cross-linking transpeptidation (inhibits cross linking of cell walls, help destabilize the cell and the cell dies – antibiotics) Archaeal Peudomurein Archaea typically do not have an outer membrane Pseudomurein: - Polysaccharide similar to peptidoglycan - Composed of two amino sugar derivatives N- acetylglucosamine acid N-acetyltalosaminuronic acid *very very similar to peptidoglycan in bacteria S-layers: protein layer supported by cell walls of many archaea ad some gram-positive bacteria - Selective barrier, cell shape, osmotic tolerance, retain proteins near the cytoplasmic membrane Gram positive vs Gram negative Gram positive: very thick peptidoglycan layer; contain teichoic acids; contain lipoteichoic acids (lipid component that anchors them to the cell membrane) Gram negative: very thin peptidoglycan layer; contain an outer membrane that contains lipopolysaccharide (LPS) - Sugars linked on the outside are the LPS – causes immune reactions (immune system recognizes LPS) - Porins: form channels through the membrane into the periplasmic space (space in between the outer membrane and in the inner membrane that is not occupied by the peptidoglycan); transmembrane protein channels that allow the diffusion of hydrophilic small molecular weight substances (non-specific channels – water filled for smaller molecules) Periplasm: space in between membranes; contains many enzymes and proteins in a gel-like matrix Outer membrane: composed of a phospholipid layer and a LPS to form the bilayer LPS: lipopolysaccharide layer; able to trigger a very strong inflammatory response in humans; membrane contains a number of proteins that provide various transport functions (lipid anchor) – endotoxin in humans O-specific polysaccharide: changes according to the bacteria Core polysaccharide: uniform regardless of the bacteria (mostly) Lipid A: toxic part ^^picture in notes Cell wall and the gram stain Structural differences in the cell wall determine the gram stain reaction. - Peptidoglycan is dehydrated by alcohol and forms and impenetrable barrier in G+ (purple stained bacteria) –the thicker the peptidoglycan is, the more impenetrable barrier, meaning the purple is trapped - G-, thin peptidoglycan and lipid-rich OM lets alcohol in and extracts purple crystal violet; counter stained pink with safranin (pink-stained bacteria) 1. Application of crystal violet 2. Application of iodine 3. Alcohol wash (decolorization) 4. Application of safranin Bacterial Cell Surface Capsule: usually a polysaccharide layer that surrounds the cell Function: attachment (bacteria loved to be attached to surfaces), protection against desiccation (don’t like to dry out), antiphagocytic (macrophages are constantly looking for pathogens to destroy, however the capsule layer hides the receptors on the membrane, camouflage to the immune system) Slime layers: similar to capsules, but are thinner and loosely attached to the cell wall Pili or fimbriae: small fibers that are used for adherence to surfaces - Very important for pathogenic microbes - Sex pilus: used in conjugation (transfer of DNA from one bacterium to another - Fimbriae often involved in attachment (biofilms) Bacterial flagella Used for locomotion Major protein is flagellin ~20,000/per filament Constructed from the inside to the outside P ring integrated into the peptidoglycan (Picture in notes is a gram negative because it has two membranes) Rotary motors: 1000 H+/rotation up to 300 rpm (electrostatic turbine) Flagellar Arrangements a. Peritrichous: multiple flagella all over (E. coli) b. Monotrichous: single flagellum c. Lophotrichous: more than one flagella at a single location Taxis: the directed movement towards and attractant or away form a repellant (towards nutrients or away from compounds) Chemotaxis: response to chemicals Aerotaxis: response to oxygen Osmotaxis: response to ionic strength Hydrotaxis: response to water Phototaxis: response to light Chemotaxis (temporal) - Run and tumble (in between run and tumbles their sensing the gradient) - net movement in direction of an attractant - Too small to sense gradient along cell - Sample environment periodically and compare to a few moments before - Temporal response, NOT spatial as they swim EUKARYOTIC CELLS Presence of a defined nucleus Nucleus: contains chromosomes; site of transcription Mitochondira: energy production Chloroplast: photosynthesis ER: protein processing; translation (rough) Golgi: protein modification and transport Vacuoles: storage or structural Lysosomes: degradation; digestion of macromolecules Peroxisome: breakdown of fatty acids Nucleus: defining organelle of eukaryal cells - Enclosed by a double phospholipid bilayer - Contains linear chromosomal DNA wound around positively charge histones (chromatin) - The site of mRNA processing - Contains nucleolus (site of ribosome assembly) - Translation occurs in the cytoplasm (not the nucleus) 1-22-16 Mitochondria and Chloroplasts Energy production – chemiosmotic process (chemical in nature and has to do with movement of ions) Contain DNA (separate from the nucleus; circular not linear), transcription and translation, and replicated independently of the host. TCA cycle in matrix (Kreb’s cycle) – mitochondria Photosynthesis occurs in thylakoid stacks – chloroplast Mitochondria – site of cellular respiration (oxidation phosphorylation) - oxidation of organic molecules used to generate a proton gradient via an electron transport chain, used to drive ATPase for ATP synthesis ATP synthase: ADPATP Amitochondirates: eukaryal microbes that do not contain mitochondria Chloroplast – use light energy to drive an electron transport chain for the generation of a proton gradient which can drive an ATPase for ATP synthesis - Flattened membrane discs are thylakoids – responsible for photosynthesis, contain chlorophyll (light reaction) - Lumen of the chloroplasts is called the strome (surrounded by IM) – site of carbon fixation (dark reaction) Protein trafficking and secretion Secretory pathway - Protein translation on rough ER - Proteins are made into the membrane of the rough ER - Transport into rough ER - Folding and modification - Transport to golgi apparatus - Modification and packaging - Transfer to plasma membrane for secretion or to another organelle Lysosomes: - Membrane-enclosed compartments - Contain digestive enzymes (hydrolysis of macromolecules) - Allow for lytic activity to occur within the cell without damaging other components - pH around 5 - residual bodies: contain indigestible material Cytoskeletal Structures Microtubules: 25nm in diameter (composed of tubulin) - motility (flagella/cilia) - chromosome segregation - movement of organelles (intracellular transport) Microfilaments: 7nm in diameter, polymers of actin - motility in pseudopodia - cell division (cytokinesis division furrow) Intermediate filaments: 8-12nm in diameter, supercoiled keratin proteins - positions organelles - nuclear structure - cell to cell interaction ALL CONTRIBUTE TO CELL SHAPE Flagella and Cilia Bacterial flagellum is very different in structure from the eukaryal flagellum. Bacterial flagellum: - tube made of protein - rotates like a boat prop - moves by proton motive force Eukaryal Flagellum: - enclosed by cytoplasmic membrane - contains microtubules (tubulin, red) - whip-like movement - Whips by hydrolysis of ATP Origin of eukaryal cells First appeared 1.6-2.1 billion years ago Arose through various endosymbiotic events. - First endosymbiotic event was the establishment of the mitrochondria - Second endosymbiotic event led to the chloroplast Bacterial origin: relation to ancient cyanobacteria and modern proteobacteria Evidence that supports the endosymbiotic theory: Lynn Margulis 1970 a. Mitochondria and chloroplast resemble bacteria in size and shape b. Membrane arrangement is consistent with the theory on how they entered the cell (double membrane) c. Membrane composition is bacteria-like in contrast to lipid composition of the plasma membrane and other organelle membranes d. DNA is found in the mitochondria and chloroplast (sequence similar to alpha-Proteobacteria and Cyanobacteria) e. Mitochondria and chloroplast divide independently of the nucleus by binary fission (bacteria-like) MICROBIAL NUTRIENT AND GROWTH All organisms need an energy for growth Energy source: phototroph (light) or chemotroph (chemical oxidation) Electron source: organotroph (electrons from organic material) or lithotroph (electrons from inorganic material) A microbe that uses light as an energy source and water as an electron source for biochemical redox reaction would be called a photolithotroph (green plants) Carob source: autotrophs (CO2) or heterotrophs (organic carbon) A microbe that used inorganic molecules for energy and an electron source and inorganic carbon for a carbon source is called a chemolithoautotroph. ^^almost all combinations are found in nature Chemical makeup of cells Macromolecules: a. Proteins - polypeptides b. Nucleic acids – DNA and RNA c. Lipids – plasma membrane d. Polysaccharides - storage and structural molecules Elements required for growth CHOPSN Carbon – major element in all classes of macromolecules Hydrogen Oxygen Phosphorus – (typically supplied as PO42-) – synthesis of nucleic acids and phospholipids Sulfur – amino acids (cysteine and methionine), vitamins (thiamine, biotin, lipoic acid) and coenzyme A Nitrogen – key element in proteins, nucleic acids Major Ions = Mg, Na, Cl, Ca, K, Fe Function: - Enzyme cofactors (required to associate with enzymes to provide function for the enzyme) - Stabilize DNA - Stabilize membranes - Enzyme activity - Osmotic balance Trace elements or micronutrients – needed in minute amounts Function: cofactors of enzymes Copper zinc, molybdenum, nickel, tungsten, selenium, manganese Iron Key component of cytochromes and FeS proteins involved in electron transport - difficult to obtain in some environments - Sliderophores: iron binding agents; used to obtain iron from insoluble mineral form Growth factors - Protoroph - Auxotroph – PURE CULTURE Containing only a single kind of microbe Contaminants: unwanted organisms in a culture Liquid or solid culture: - Solid media are prepared by addition of agar - When grown on solid media, cells form isolated masses (colonies) Culture Media Nutrient solutions used to grow microbes in the laboratory 1. Defined media: precise chemical composition is known 2. Complex media: composes of digests of chemically undefined substances Media can additionally be: Selective Media: contains compounds that selectively inhibit growth of some microbes but not others Differential Media: contains an indicator, usually a dye that detects particular chemical reactions occurring during growth. Allows researcher to distinguish between different bacteria (you can have selective defined media or selective complex media or differential defined media or a selective defined media) Cardinal temperatures of all microbes Can be a very steep drop off from optimum temperature to lethal. Archaea can grow much hotter than bacteria. (hyperthermophilic archaea) Eukaryotes stop existing above 60 degrees Celsius. Acidity and Alkalinity – pH The internal pH of a cell must stay relatively close to neutral even though the outside pH is highly acidic or basic. Osmotic effects Water availability = water activity: vapor pressor of air over a solution compared to vapor pressure over H2O. Values 0-1. Closer to 0, more salty. Solution with a high salt or sugar concentration = low water activity. Osmosis: water diffusion across a biological membrane Water moves from areas of low solute concentration to areas of high solute concentration. Hypertonic – high concentration of dissolved components outside; shrinking cell walls (plasmolysis) Water tends to move into cells (hypotonic, positive water balance) unless the solute concentration outside is greater than inside (hypertonic, low water activity) Cells fight plasmolysis in hypertonic solutions by: a. Pumping solutes in; pump in salts from the environment to equalize inside and outside solute concentrations b. Producing compatible solutes to raise their internal solute concentration Halophiles: salt loving organisms; have an absolute requirement for saline conditions - Their enzymes and ribosomes require high salt concentrations; not active at lower concentrations Besides halophiles: a. Osmophiles: organisms that live in an environment with high sugar concentrations b. Xerophiles: organisms able to grow in very dry environments Osmotolerant or Halotolerant – an organisms that will grow over a wide range of water activity (sugar or salt as the solute) OXYGEN Thioglycolate broth: complex medium that separates microbes based on oxygen requirements; reacts with oxygen so oxygen can only penetrate the top of the tube. a. obligate aerobe - need oxygen b. Obligate anaerobe - cannot have oxygen c. Facultative - can us oxygen if present and will grow without oxygen d. Micro-aerophile - use only low levels of oxygen) e. Aerotolerant Anaerobe - oxygen is OK but cannot use Why is oxygen problematic? Toxic oxygen species. Toxic oxygen needs to be detoxified: catalase test - adding cells to H2O2 - if it bubbles, it is positive for catalase - production of oxygen Superoxide dismutase is indispensable in obligate aerobes. Superoxide reductase – used by some anaerobes – no O2 production Most obligate anaerobic bacteria do not contain these enzymes – this may contribute to their inability to tolerate oxygen. Growing anaerobic prokaryotes - anaerobic jar: any hydrogen and oxygen is converted to water by the palladium catalysts pellets - anaerobic chamber BACTERIAL CELL DIVISION Binary fission 1 generation time: 2 cells to one cell Bacterial cytoskeleton function in cell division, shape, and chromosome segregation. a. FtsZ protein: tubulin-like protein functions in cell division through the formation of the Z-ring b. FtsA protein: actin-like protein that functions to anchor FtsZ to the cytoplasmic membrane and recruitment of other division proteins c. Min proteins: directs formation of FtsZ ring and divisome complex to the division plane d. MreB protein: actin-like protein that functions in cell wall assemble to give the cell shape (in rods) Divisome: ring-like structure of division protein, mainly Fts proteins. Main protein FtsZ anchored to cell membrane by FtsZ and ZipA. Min system: two helical protein MinCD and MinE; ensures division is symmetrical; inhibits divisome formation at the poles (because they spend so much time there) MinE can spend more time at the middle of the cell and can therefore recruit FtsZ at mid-plane.
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