Exam 2 Bacteriology

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Characteristics of LPS

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Frequently toxic to animals, toxic portion is lipid A, heat stable; induces fever, shock, diarrhea, and vomiting in humans, referred to as an endotoxin because it is generally embedded within the cell surface and released in large amounts only when the cells lyse

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Three components of LPS

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O-specific polysaccharide (O antigen), core polysaccharide, lipid a

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O-antigen

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Part of LPS; highly variable between species and strains; used to type strains

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Lipid A

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Component of LPS; fatty acids linked to NAG disaccharide (part of peptidoglycan) that anchors the LPS within the outer membrane

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Superantigen

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Type I protein exotoxin; toxin that nonspecifically activates large populations of T-cells to produce cytokines; toxic shock syndrome toxin made by S. aureus; binds indiscriminately to MHC-II on APCs and to the TCRs on T-cells; IL-2 released by T-cells csudrs nausea, vomiting, and fever

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Type I protein exotoxins

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Toxins that nonspecifically activate large populations of T-cells to produce cytokines

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Type II protein exotoxins

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Pore forming toxins

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What are the roles of pore forming toxins

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Release of nutrients/cell death (S. aureus produces alpha hemolysin), delivery of effectors to cytoplasm (streptolysin O), lysis and escape from the phagocytic vaculoe (listeriolysin O)

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Alpha-hemolysin

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Produced and secreted as monomers, diffuse towards target membranes, bind to target membrane (target membranes are lipids, sugars, proteins), after binding, the monomers diffuse until enough come together to form an oligomeric ring (7-50 monomers may be required to form a pore) which inserts spontaneously into the membrane

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Examples of cholesterol-dependent toxins and functions

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Lisyeriolysin O and streptolysin O; bind specifically to cholesterol in the cell membrane

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Examples of RTX toxins

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HyA, hemolysin found in Vibrio cholera

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Phospholipases

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Enzymes that cleave phospholipids, disrupting the host cell membranes

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Examples of type II exotoxins

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Alpha-hemolysin, cholesterol-dependent toxins, RTX toxins, phospholipids

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Type III protein exotoxins

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The A:B family of toxins

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What does the A:B stand for

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A: catalytic Activity that's encoded by the A domain

B: Binding domain that's encoded by the B domain

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What does the activation of exotoxin A require

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Proteolytic processing and disulfide bond reduction

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What is the biochemical activity of some A:B toxins

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ADP-ribosyltransferase activity and proteases

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What is ADP-ribosylation of elongation factor 2 and name examples

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Inhibits protein translation in eukaryotic cells

Examples include exotoxin A in P. aeruginosa

Diptheria toxin in C. diphtheria

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What do proteases do in botulinum and tetanus toxins

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Botulinum: blocks release of acetylcholine (neurotransmitter) in peripheral nerve endings which results in flaccid paralysis

Tetanus: blocks neurotransmitter in the CNS which results in spacid paralysis or lock jaw

Both: prevent the fusion of secretory vesicles with the synapse by cleaving SNARE and SNAP proteins

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Sec system

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The general secretory system

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Two classes of proteins translocated by sec system

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Secretory proteins and integral inner membrane proteins

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Where can secretory proteins traffic to

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Periplasm, OM, be completely released from cells

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What types of bacteria is the sec system found in

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Gram negative and gram positive bacteria

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What percentage of proteins go through Sev

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20%

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What are signal peptides composed of and what do thy do

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Charged region, hydrophobic core cleavage site (30 amino acids long) and the mature domain

Secretory proteins are targeted via signal peptides

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What does SecYEG do

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Forms a translocation pore through which a protein is secreted in an unfolded conformation; the protein will adopt a final confirmation in the periplasm

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What is SecYEG

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A translocase

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SecA

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ATPase

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SecB

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Maintains preproteins in exports competent conformation and prevents premature folding and aggregation

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LPase

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Leader peptidase

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Type II secretion system

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The EPS system; used to secrete both the A and B portions of the cholera toxin out OG the bacteria; spans the OM and IM in Vibrio cholera

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The EPS system

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Type II secretion system involved in cholera toxin secretion

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Type V secretion system

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Autotransporters; a sec dependent secretion system; the protein is exported as an extended linear chain of amino acids (polypeptide)

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Composition of an auto transporter

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Three domains: signal peptide, passenger domain, beta-domain; the secretion pore required for transport across the OM is contained within the precursor of the secreted protein itself; the beta-domain spontaneously inserts into the OM in a beta-barrel conformation (multiple amphipathic antiparallel beta sheets), and processing of the passenger domain can be done by the passenger domain itself, OM proteases or sometimes they remain surface bound

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Example of type V secretion system

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IgA proteases from H. influenza

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TAT system

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The twin-arginine transport system

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What types of bacteria is the TAT system found in

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Both gram positive and gram negative bacteria

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What does the TAT system do

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Transports proteins with double-arginine in their signal sequence

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What are functions of the Tat translocase pathway

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Translocation of co-factor bound proteins to the periplasm or IM, translocation of multi-subunit enzymes, translocation of proteins which are tightly bound and therefore incompatible with the Sec translocase, and secretion of virulence determinants

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What is the composition of the TAT translocase and how does it work

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Consists of TatABC that forms a large pore complex and works when the proton motive force across the membrane energized the transport step (an electrochemical gradient)

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Sec-dependent vs tat-dependent secretory pathways

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Sec-dependent only transports unfolded proteins and works a classic ss pathway and the tat-dependent system transports folded cofactor bound and works a twin-arginine ss

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Name sec-dependent secretion systems

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Type II and type V secretion systems

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Name sec-independent secretion systems

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Type I, type IV, type VI secretion systems

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Type I secretion system

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E. coli's alpha hemolysin, has two components in the IM and one in the IM; HlyB couples ATP hydrolysis to export HlyB and D

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HlyB

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Type I, an ABC transporter

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HlyD

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Type I, a membrane fusion protein

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TolC

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An OM protein involved in many export pathways, also involved in drug efflux pump

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HlyA

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Carries a C-terminal secretion signal, binds HlyB and induces the interaction of HlyD with TolC

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Type III secretion system

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Used to inject bacterial toxins directly into mammalian cells, usually contact dependent

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Effector proteins

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Bacterial toxins

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Type IV secretion system

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Conjugation system adapted to deliver effector molecules to host cells, transfers DNA from one cell to another, used to take up or release DNA

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Type VI secretion system

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Requires set of 15 conserved genes, effects of toxin are contact dependent, used against eukaryotic cells and other bacteria, phage-related extracellular components