Summaries – 1 BI-311

Ferdinand Cohn Founded the field of bacteriology Recognized distinction between prokaryotic and eukaryotic cellular organization Discovered bacterial endospores The Historical Roots of Microbiology: The Science

The Historical Roots of Microbiology: Louis Pasteur Discredited the theory of Spontaneous Generation. Introduced control of microbial growth. Discovered lactic acid bacteria Role of yeast in alcohol fermentation Rabies vaccine

The Historical Roots of Microbiology: Robert Koch Growth of pure cultures of microorganisms Solid growth media Discovered cause of tuberculosis. Developed criteria for the study of infectious microorganisms Kochst Postulates.

Koch’s Postulates OBSERVE: The presence of suspected pathogenic microorganism correlates positively with the symptoms of the diseased and negative with healthy control ISOLATE the suspected pathogen into axenic culture INFECT a healthy animal with cultured strain. Observe whether the same symptoms show RE-ISOLATE the pathogen from the new victim and compare both cultures

The Historical Roots of Microbiology: General Microbiology - Microbial Ecology and Diversity Martinus Beijerinck Enrichment Culture Technique Concept of Virus Sergey Winogradsky Concept of Chemolithotrophy and Autotrophy

Monera Protoctista fungi Animals plants EUKARYOTES PROKARYOTES Five Kingdoms “Crown species”

Incident light microscopy (dissecting) Transmitted light microscopy (compound) Phase contrast Dark field Differential Interference Contrast (DIC) Fluorescence microscopy Confocal Scanning Light Microcopy (CSLM), Transmission electron microscopy (TEM) Scanning electron microscopy (SEM) The atomic force microscope

Differential Interference Contrast (DIC) and Confocal Scanning Light Microcopy (CSLM) allow for greater three-dimensional imaging than other forms of light microscopy, Confocal microscopy allows imaging through thick specimens. The atomic force microscope yields a detailed three-dimensional image of live preparations.

Electron microscopes use electron beams instead of light. They have far greater resolving power than do light microscopes, the limits of resolution being about 0.2 nm. Two major types of electron microscopy are performed: Transmission Electron Microscopy (TEM), for observing internal cell structure down to the molecular level, and Scanning Electron Microscopy (SEM), useful for three-dimensional imaging and for examining surfaces.

Scanning Electron Microscopy – SEM Glutaraldehyde-fixed, critical point-dried, gold- paladium coated

Eukaryotic cell Freeze-etched preparation Carbon-coated, Gold-shaded, TEM image

Macromolecule s Organic chemistry = chemistry of carbon Biochemistry = chemistry of macromolecules Water = solvent & chemical bonding properties: polarity, hidrophilic vs. hydrophobic H-bonds, glycosidic, esteric, etheric, peptide. Biogenic elements = C, O, H, N, S, P construct polymers from monomers: polysaccharides, (phospho-)lipids, polypeptides, polynucleotides

CARBOXYL ALDEHYDE ALCOHOL KETO ESTER PHOSPHO-ESTER THIOESTER ETHER ACID ANHYDRIDE PHOSPHO ANHYDRIDE

The cell walls of Bacteria contain a polysaccharide called peptidoglycan. This material consists of strands of alternating repeats of N-acetylglucosamine and N- acetylmuramic acid, with the latter cross-linked between strands by short peptides. Many sheets of peptidoglycan can be present, depending on the organism. Archaea lack peptidoglycan but contain walls made of other polysaccharides or of protein. The enzyme lysozyme destroys peptidoglycan, leading to cell lysis in Bacteria but not in Archaea

In addition to peptidoglycan, gram-negative Bacteria contain an outer membrane consisting of lipopolysaccharide, protein, and lipoprotein. Proteins called porins allow for permeability across the outer membrane. The space between the membranes is the periplasm, which contains various proteins involved in important cellular functions.

Prokaryotic cells often contain various surface structures. These include: fimbriae pili S-layers capsules slime layers. These structures have several functions, but a key one is in attaching cells to a solid surface.

Prokaryotic cells often contain internal granules such as sulfur, PHB, polyphosphate, PHAs, and magnetosomes. These substances function as storage materials or in magnetotaxis.

Gas vesicles are small gas-filled structures made of protein that function to confer buoyancy on cells. Gas vesicles contain two different proteins arranged to form a gas permeable, but watertight structure: Gas Vesicle Proteins GVP-a and GVP-c.

The endospore is a highly resistant differentiated bacterial cell produced by certain gram-positive Bacteria. Endospore formation leads to a highly dehydrated structure that contains essential macromolecules and a variety of substances such as calcium dipicolinate and small acid- soluble proteins, absent from vegetative cells. Endospores can remain dormant indefinitely but germinate quickly when the appropriate trigger is applied.

Motility in most microorganisms is due to flagella. In prokaryotes the flagellum is a complex structure made of several proteins. Most of these proteins are anchored in the cell wall and cytoplasmic membrane. The flagellum filament, which is made of a single kind of protein, rotates at the expense of the proton motive force, which drives the flagellar motor.

Prokaryotes that move by gliding motility do not employ rotating flagella, but instead creep along a solid surface by any of several possible mechanisms.

Motile bacteria can respond to chemical and physical gradients in their environment. In the processes of chemotaxis and phototaxis, random movement of a prokaryotic cell can be biased either toward or away from a stimulus by controlling the degree to which runs or tumbles occur. The latter are controlled by the direction of rotation of the flagellum, which in turn is controlled by a network of sensory and response proteins.

Microbial Metabolism Biocatalysis & Energy Generation Phosphorylation Oxidation & Reduction Fermentation & Respiration Chemiosmosis: Proton Motive Force ATPase Motor Energy Yielding Metabolic Systems Biosynthesis

∆G 0 ' versus ∆G standard conditions pH 7, 25°C

The chemical reactions of the cell are accompanied by changes in energy, measured in kilojoules (kJ). A chemical reaction can occur with the release of free energy (exergonic) or with the consumption of free energy (endergonic). 1 calorie = Joules

Energy G 0’ f = free Energy of formation for elements G 0’ f = 0 ΔG 0’ = change in free Energy in reactions ΔG 0’ of the reaction: A+B  C+D equals ΔG 0’ [C+D] - ΔG 0’ [A+B] products - reactants if +, the reaction is ENDERGONIC if -, the reaction is EXERGONIC ΔG 0’ does not affect the rates of reaction

The reactants in a chemical reaction must first be activated before the reaction can take place, and this requires a catalyst. Enzymes are catalytic proteins that speed up the rate of biochemical reactions. Enzymes are highly specific in the reactions they catalyze, and this specificity resides in the three- dimensional structure of the polypeptide(s) in the protein.

Enzyme Biocatalysis Specific substrate binding Substrate orientation o active sites Lowering the activation energy

The energy released in redox reactions is conserved in the formation of certain compounds that contain energy-rich phosphate or sulfur bonds. The most common of these compounds is ATP, the prime energy carrier in the cell. Long-term storage of energy is linked to the formation of polymers, which can be consumed to yield ATP.

Microbial Metabolism Biocatalysis & Energy Generation Phosphorylation Oxidation & Reduction Fermentation & Respiration Chemiosmosis: Proton Motive Force ATPase Motor Energy Yielding Metabolic Systems Biosynthesis

Oxidation–reduction reactions involve the transfer of electrons from electron donor to electron acceptor. The tendency of a compound to accept or release electrons is expressed quantitatively by its reduction potential, E0’.

The transfer of electrons from donor to acceptor in a cell typically involves one or more electron carriers. Some electron carriers are membrane-bound, whereas others, such as NAD+/NADH, are freely diffusible, transferring electrons from one place to another in the cell.

The energy released in redox reactions is conserved in the formation of certain compounds that contain energy-rich phosphate or sulfur bonds. The most common of these compounds is ATP, the prime energy carrier in the cell. Long-term storage of energy is linked to the formation of polymers, which can be consumed to yield ATP.

Fermentation and respiration are the two means by which chemo- organotrophs conserve energy from the oxidation of organic compounds. During these catabolic reactions, ATP synthesis occurs by way of either substrate-level phosphorylation (fermentation) or oxidative phosphorylation (respiration).

Glycolysis is a major pathway of fermentation and is a widespread means of anaerobic metabolism. The end result of glycolysis is the release of a small amount of energy that is conserved as ATP and the production of fermentation products. For each glucose consumed in glycolysis, 2 ATPs are produced.

Respiration involves the complete oxidation of an organic compound with much greater energy release than during fermentation. The citric acid cycle plays a major role in the respiration of organic compounds.

When electrons are transported through an electron transport chain, protons are extruded to the outside of the membrane forming the proton motive force. Key electron carriers include flavins, quinones, the cytochrome bc1 complex, and other cytochromes, depending on the organism. The cell uses the proton motive force to make ATP through the activity of ATPase.

Chemo – Energy from chemical reactions Organo – – trophic of organic compounds feeding Hetero – Carbon from organic sources

Electron acceptors other than O 2 can function as terminal electron acceptors for energy generation. Because O 2 is absent under these conditions, the process is called anaerobic respiration. Chemolithotrophs use inorganic compounds as electron donors, while phototrophs use light to form a proton motive force. The proton motive force is involved in all forms of respiration and photosynthesis.

Energy from: Chemical reactions or Light Chemo- Photo- of: inorganic or organic compounds Litho- Organo- Source of carbon : CO 2 or (CH 2 O) n Auto- Hetero-

Amino acids are formed from carbon skeletons generated during catabolism while nucleotides are biosynthesized using carbon from several sources.

Lipids Fatty acids are synthesized two carbons at a time and then attached to glycerol to form lipids.