Microbial structure (2)

BC Yang

Structural approach : Is it a view of artifact?

For lecture only

Shapes

Bacillus

Coccus

Coccobacillus

Fusiform bacillus

Spirillum

Spirochete

Vibrio

Staphylococcus aureus

Pseudomonas aeruginosa

Bacillus anthracis

Vibrio cholerae

Fusiform bacilli

Spirillum volutans

Borrelia burgdorferi

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Structural view of bacteria (1)

Microscopic prokaryotes (no nucleus nor membrane-bound organelles)

Contain ribosomes Enfolding of the cell membrane carry on

photosynthesis & respiration Surrounded by protective cell wall containing

peptidoglycan (protein-carbohydrate) Many are surrounded by a sticky, protective coating

of sugars called the capsule or glycocalyx

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Structural view of bacteria (2)

One circular chromosome and some small DNA called plasmids

May have short, hairlike projections called pili on cell wall to attach to host or another bacteria when transferring genetic material

Some can move by flagella, gliding over slime they secrete ( e.g. Myxobacteria)

Some can form protective endospores around the DNA when conditions become unfavorable

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Detail Structure

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Structure and functionSTRUCTURE FUNCTIONCell Wall protects the cell and gives shape (5-20 atm resistant)

Outer Membraneprotects the cell against some antibiotics (only present in

Gram negative cells)

Cell Membraneregulates movement of materials into and out of the cell;

enzymes of respiration

Cytoplasm contains DNA, ribosomes, and organic compounds

Chromosome carries genetic information inherited from past generations

Plasmidcontains some genes obtain through genetic

recombination

Capsule, and slime layer

protects the cell (immune attack) and assist in attaching the cell to other surfaces

Endosporeprotects the cell against harsh environmental conditions,

(heat or drought)

Pilus (Pili) attaching to other surfaces (for genetic recombination)

Flagellum moves the cell

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Differences

Eukaryote ProkaryoteMajor groups Algae, fungi, protozoa, plant,

animalbacteria

Size >5 m 0.5-3m

Nuclear structures

Nucleus Classic membrane No nuclear membrane

Chromosome Diploid genome; multiple DNA Haploid, single DNA

Cytoplamic structures

Mitochondria Present Absent

Golgi bodies Present Absent

ER Present Absent

Ribosome 80S(60S+40S) 70S(50S+30S)

Cytoplamic membrane Contains sterols No sterols

Cell wall Absent or with chitin, cellulose + lipid, peptidoglycans

Reproduction Sexual and asexual Fission

Movement Complex flagellum (9+2) Simple flagellum

Respiration Via mitochondria On cytoplamic membrane

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Cell envelope: Cytoplamic membrane+ cell wall+ capsules + loss slime etc….

Gram-positive cells: Cytoplamic membrane+

thick peptidoglycan layer + capsules

Gram-negative cells: Cytoplamic inner

membrane+ thin peptidoglycan layer + periplamic space + outer membrane + + capsules /loss slime

BC Yang

high magnification AFM image of the surface of a single Pseudomonas putida

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• The main component of the bacterial cell wall is peptidoglycan, a hydrated, semi-rigid polymer of two sugar derivatives: N-Acteylglucosamine (G) and N-Acetylmuramic acid (M). Bound to the sugar M are amino acids: Alanine -- Glutamic acid -- Meso-diaminopimelic acid (DAP) (Gm-) or Lysine (Gm+) -- Alanine.

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Glucosamine?

Teichoic acidAre found exclusively in gram positive organisms. Are formed as polymers of glycerol or ribitol through phosphodiester linkages.

http://www.cvm.uiuc.edu/courses/vp331/Structures_in_pathogenesi1.htmlBC YangFor lecture only

A very complex net

http://www.arches.uga.edu/~kristenc/cellwall.html

Synthesis of cell wall: The assembly of the wall components begins with the synthesis of precursors in the cytoplasm, their transport across the cell membrane, and their final polymerization. Eventually, penicillin-binding proteins catalyze covalent reactions that result in the extension, cross-linking between glycan strand, morphogenessis and eventual separation of the murein sacculus.

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透過骨架來改變形狀

Fluorescent image of the CreS-GFP/CreS hybrid strain

BC Yang

1. Crescentin : creS2. 430-residues protein3. Distinct 7-residue repetitive pattern4. coiled-coils: many fibrous proteins in eukaryotes

Look into the detail

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Gram positive in Gm+ organisms, an interbridge exists between the Lysine

molecule extending from one sugar backbone to the terminal Alanine of the second.

Gm+ organisms have cell walls that may contain as much as 90% peptidoglycan, with membrane associated proteins and Teichoic acids comprising the remaining components.

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Gram negative cross-linking of peptidoglycan by bond between the DAP molecule

extending from one sugar backbone and the terminal Alanine of another. typically composed of only about 10% peptidoglycan and possess an outer

membrane that houses the membrane associated proteins and LPS. The peptidoglycan lies in-between the periplasm.

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The acid-fast cell wall of Mycobacterium contains peptidoglycan, and a large amount of glycolipids such as

mycolic acid, arabinogalactan-lipid comlex, and lipoarabinomannan.

Large-scale molecular dynamic simulation of a lipopolysaccharide membrane solvated in a 4.2 nm water box.

Lipopolysaccharides is an amphiphile composed of three regions: O-polysaccharide (the O- or somatic-antigen), the core polysaccharide and lipid A. Lipid A is anchored in the outer membrane. LPS is also known as endotoxin.

BC Yang

lipid

saccharide

For lecture only

Endotoxins are thermostable, lipopolysaccharide components from the cell walls of viable or nonviable gram-negative microorganisms.

Measured endotoxin will include endotoxin that was derived from: 1) Any living cells that are present, 2) Cell wall debris from dead or dying cells, and 3) Outer membrane fragments that are released during cell growth.

An endotoxin unit, EU, is equivalent to approximately 200 pico-grams of endotoxin.

The LAL (Limulus Amebocyte Lysate) test for endotoxins can be sensitive down to 0.03 EU/ml

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鱟

Not confuse with exotoxin

BC Yang

(a) The x-ray crystal structure for the tetanus toxin showing how the amino acid chain is folded and (b) its calculated molecular surface showing sites 1 and 2, predicted binding sites for ligands.

Tetanus toxin:

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Cytoplasmic membrane

5-10 nm unit membrane Absence of sterols, except mycoplasma Semifluid state, form mesosomes Selective permeability and transport Electron transport /oxidative ATP generation Bioxynthesis

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Phospholipids Major components of the cell

membrane Consists of two fatty acids and the

third hydroxyl group of glycerol is joined to a phosphate group

being amphipathic, having both a hydrophobic (hydrocarbons tails) and a hydrophilic region (phosphate head)

In water, they self-assemble into aggregates so that the phosphate heads make contact with the water and the hydrophobic hydrocarbon tails are restricted to water-free areas (micelle or phospholipid bilayer)

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How to get across the polar barrel

Bacterial membrane proteins - OmpA (left) and GlpF (right).

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ATP-dependent secretion systems

There are five major secretion systems.

Type I and type III secrete proteins across both the inner membrane

and the cell envelope (outer membrane) in one step; secreted proteins

do not make an intermediate stop in the periplasm. Type I systems

are composed of far fewer components than type III systems.

Type II and type III systems share a similar cell envelope component.

The type III secretion system of Gram-negative bacterial pathogens

injects virulence factors into host cells. The needle complex is drawn

on the basis of its appearance in electron-microscopy images (see

inset), with several ring structures that span the inner and outer

membranes. Very similar to flagella system

For lecture only

Bacterial secretion systems: type I, type II, and type III.

http://www.cdc.gov/ncidod/eid/vol2no4/mecsas.htm

For lecture only

The known type IV systems differ with respect to the route of substrate translocation. The A. tumefaciens T-DNA transfer system and the H. pylori CagA system are thought to export substrates in one step across the membrane directly to the eukaryotic cytosol. The B. pertussis Ptl system is thought to export PT in two steps across the cell envelope to the extracellular milieu. Secreted holotoxin then binds to the mammalian cell membrane.

http://www.blackwell-synergy.com/links/doi/10.1046/j.1365-2958.2001.02302.x/full/For lecture only BC Yang

Bacterial chromosome No nuclear membrane

Small polyamines and magnesium, histone-like

Lack of mitotic apparatus; fission

~3 x 109bp (1mm long), supercoiled state, single haploid

Circular (most bacteria) or linear (Streptomyes sp.)

Associated with mesosomeDNA spread of E. coli

BC YangplasmidFor lecture only

may stay inactive several years, then re-activate when conditions favorable

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Calcium bound to dipicolinic acidKeratin-like protein coat

Ca+2

Spores

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1. the bacterium senses that its home or habitat is turning bad2. it makes a copy of its chromosome3. the rubbery cell membrane that surrounds the bacterial cell fluid

begins pinching inward around this chromosome copy. 4. the membrane of the mother cell surrounds and swallows up the

daughter cell.5. between these two membranes a thick wall forms made out of

stuff called peptidoglycan.6. a tough outer coating made up of a bunch of proteins forms

around all this, closing off the entire daughter cell.

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Capsule Bacterial capsules outlined by India

ink viewed by light microscopy. A discrete layer of polysaccharide

surrounding the cells. Sometimes bacterial cells are

embedded more randomly in a polysaccharide matrix called a slime layer or biofilm.

Polysaccharide films that may inevitably be present on the surfaces of bacterial cells, but which cannot be detected visually, are called glycocalyx.  

BC Yang

Capsule Stain of B. anthracis:

Capsule appears pink, bacterial cells purple

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Capsule

Composed of poly-(D-glutamic acid), single antigenic type + proteins

Nontoxic, serves as an impedin in establishment of infection

Production enhanced in the presence of Na+-bicarbonate

may be plasmid-borne

prevents desiccation of the bacteria prevent phagocytosis by larger microorganisms and the

white blood cells of invaded host organisms help the bacteria adhere to the host substrate assist in warding off attacks by phages

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Pili (Fimbriae) singular pillus helical filaments, with ~1000 copies of

the major pilin, plus one or a few copies of ~5 minor pilins. The minor proteins provide binding specificity, membrane anchoring, and adapter functions.Mu, Egelman, Bullitt, 2002. J. Bact. 284(17):4868-4874Bullitt, & Makowski, 1995. Nature 373(6510):164-167

a) Adhesionb) In pathogenesisc) In environmentd) Sex pili

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Sex pili also called as type IV secretion system!

Flagellasingular flagellum

A tiny motor machine

BC YangFor lecture onlyShare similarity with type III secretion system

Each flagellum is a rigid structure The basal body causes rotation of the structure -- like a

propeller. The rate is about 12000 rpm Energy is derived from a proton gradient. Power has been

calculate to be about 10-15 amperes of current. Average velocities are in the range of 20 - 80 µM/s. About 1/2

foot per hour. A cheetah is 4 feet long and moves about 70

mph. 25 body lengths/second. A microbe that is 2 µM long,

moves 10 to 40 body lengths per second. (some bacteria are as

mobile as higher animals) Movement is a bias random walk during chemotaxis

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Swing and tumble

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