Also: Complete Genome Sequence of the Methanogenic Archaeon, Methanococcus jannaschii

Bult et al. Science 273: 1058-1073

See also pages 1043-1045 of same issue

ENVELOPE (Overview comparison given by Dr. Bernlohr; compare Fig. 1.12 and Fig. 2.1)

CELL MEMBRANE

CELL WALL

Gram Positive (G+)

Acid-Fast Cell Wall

Gram Negative (G-)

Periplasm

Crystalline Surface Layer

CAPSULE

FLAGELLA

PILI

POLYSOMES

STORAGE GRANULES / INCLUSION BODIES

Bilayer = "Unit" membrane 8 nm thick (hydrophobic core is ~4 nm thick)

Eubacteria: phospholipid bilayer consisting of 70% protein + 30% phospholipids

E. coli: phosphatidylethanolamine (75%)

phosphatidylserine (2%)

phosphatidylglycerol (18%)

cardiolipin (glyercol-linked dimer of phosphatidylglycerol) (5%)

For structures, see p. 15 of text

G- bacteria, mostly phospholipids

G+glycolipids (sugar-glycerol-(fatty acid)2) not uncommon; may replace sterols, provide stability to membrane.

Sterols absent (except for mycoplasmas), but distantly related poly-cyclic poly-terpenoids known as HOPANES are found in many species.

Membrane represents a two-dimensional fluid at temperatures near the growth temperature.

Fatty Acids: 43% Palmitic Acid (16:0)

33% Palmitoleic Acid (16:1)

25% cis-Vaccenic Acid (18:1)

CH₃ (CH₂)_n-COOH

Trends: Organisms that grow at higher temperatures have more saturated fatty acids and longer chains

Organisms that grow at lower temperatures have fatty acids with shorter chain lengths and more poly-unsaturated fatty acids. This keeps membranes fluid even at temperatures below freezing. Seems to be important to the functioning of membrane proteins, especially permeases.

Eubacteria and Eucarya have ESTER-LINKED fatty acids in lipids

Archaea have ETHER-LINKED lipids with C₂₀-C₄₀ ISOPRENOID (branched hydrocarbons) alcohols. Because of head and tail group variability, archaea have more variable lipid composition (5-25 different species of lipids in a single organism). Ether-linked lipids probably more stable in harsh environments, although not an absolute correlation)

1. Osmotic Barrier

2. Transport of specific nutrients and ions

3. Synthesis of lipids

4. Synthesis of murein (peptidoglycan)

5. Assembly and secretion of envelope proteins

6. Respiratory electron transport

7. Chromosome segregation

8. Environmental sensing (e.g., chemotaxis)

Wall prevents cell from osmotic lysis, chemical and physical barrier to the outside world. The structure of the peptidoglycan layer gives cell its characteristic shape. It is a single molecule--very strong!

Mostly PEPTIDOGLYCAN with some TEICHOIC ACIDS interspersed

Much higher degree of cross-linking in G+ bacteria than in G- bacteria (cross-linking can be 75-100% in G+, but as low as 25-30% in E. coli).

Sugar polymers: (N-acetylglucosamine and N-acetylmuramic acid co-polymer)

Peptide chains: tetrapeptide chain (L-Ala-D-Glu-m-DAP-D-Ala). DAP can be replaced by several other di-amino amino acids

Cross Bridges: Not present in most G- bacteria, but common in G+. Example: Staph. aureus has a pentaglycine (Gly₅) bridge with nearly 100% cross-linking efficiency.

Variable in G+ walls, but can account for up to 50% of mass of the wall. Highly antigenic and useful in identifying specific species/strains of bacteria. Usually covalently linked to glycan chains of PG

TEICHOIC ACIDS are polymers of glycerol phosphate or ribitol phosphate with attached sugars or amino acids.

TEICHURONIC ACIDS are acidic polysaccharides containing uronic acids (e.g., glucuronic acid) and other sugars

Function still unknown. Some have attached lipids (LIPOTEICHOIC ACIDS) and may serve to anchor the wall to the cell membrane. In the pathogen Strep. pyogenes, lipoteichoic acids associate with "M-PROTEIN" forming long fibrils that facilitate attachment to animal cell surfaces. Possibly functionally analagous glycolipids found in other G+ bacteria

LYSOZYME can convert G+ bacteria into PROTOPLASTS, if isoosmotic medium is used (e.g., 0.5 M sucrose or ~0.15 M NaCl)

Mycobacteria (causative agents of tuberculosis and leprosy) contain waxy lipids known as MYCOLIC ACIDS. These are long-chain hydrocarbons (C₂₄-C₆₀) substituted with sugars and other groups.

Protection against hydrophobic compounds and acids. Allows specific staining method, known as acid-fast staining. Organisms are resistant to killing action of white blood cells, and can in fact live inside the phagocytic vesicles of such cells.

Nutrient uptake may be limited by the waxy layer; mycobacteria grow rather slowly

Totally different solution to the same problem. Murein/PG layer is much thinner, and is probably a monolayer in E. coli.

Peptidoglycan is probably synthesized by insertion of rings (about 200 growing sites per cell; it takes about 1100 rings to surround the cell). Spacing of rings is about 1.25 nm

OUTER MEMBRANE is chemically distinct from the cytoplasmic membrane; unusually resistant to chemicals and hydrophobic compounds,including many antibiotics. Inner leaflet similar to other membranes (phospholipids); outer leaflet is made up of LIPOPOLYSACCHARIDE (LPS), a complex and biologically unique molecule. Acts as ENDOTOXIN, producing symptoms such as fever, shock, and hemorrhage.

LPS consists of 3 portions (SEE p. 17 of text): LIPID A (2 glucoseamines + phosphates with C₁₄ 3-OH myristic acid (unique fatty acid)

Core: short branched sugar chain, relatively constant in all G- bacteria; contains two characteristic sugars: one, a HEPTOSE, the other, KDO, or KETO-DEOXYOCTONOIC ACID

O-ANTIGEN: Long carbohydrate chain, up to 40 sugars in length; cover the bacterial surface; very effective in excluding hydrophobic compounds. E. coli O157:H7 specifies a specific O-antigen carrying strain of E. coli that is usually enterohemorrhagic (causes bleeding of the intestinal lining due to Shiga-like toxin)

This LPS "monomer" usually is covalently joined into trimeric units (on average) through pyrophosphate linkages to the sugars of lipid A; there are probably covalently linkages to proteins as well.

Murein (Braun's) lipoprotein: most abundant protein of outer membrane; 700,000 copies per cell in E. coli; anchors PG to outer membrane; is covalently linked to PG via DAP and has attached fatty acids for interaction with outer membrane. Stabilizes cell surface

The lipid bilayer nature of the outer membrane also blocks entry of hydrophilic compounds however.

Solution: PORINS. These are proteins, usually trimeric, that form passive diffusion channels through the outer membrane. Allow molecules of ~600-700 Da to pass, but not any macromolecules. Exact composition determined by environmental conditions, including osmotic pressure and barometric pressure

Porins differ in number and type of channels. 3 types. 1. OmpF-like: 3 channels join to form a single channel; 2. 3 completely independent channels; 3. "Filter type" with a single, constricted and often selective passage.

Specific carriers required for some hydrophilic compounds (e.g., vitamin B₁₂ or Fe⁺⁺-chelator complexes).

Regions (~200 per cell) where cytoplasmic membrane and outer membrane make contact. Not much known about them. Transient? Permanent? Function? Also, Periseptal Annuli--a ring of adhesion near cell division septum

Lysozyme can not reach peptidoglycan in untreated cells. Freeze-thaw or treatment with chelators disrupts OM, allows protein to have access to PG. Blocking PG synthesis with penicillin also works.

20-40% of the volume of a G- cell lies between CM and OM; possible osmoregulation by membrane-derived oligosaccharides.

Things found in Periplasmic Space:

Binding Proteins: for amino acids, sugars, vitamins, ions

Degradative enzymes: phosphatases, proteases, endonucleases

Detoxifying enzymes: b-lactamase (penicillinase)

Biosynthesis: peptidoglycan biosynthesis

Energy production: cytochromes,

Environmental sensing: chemical sensors

Although G+ bacteria don't have OM, they still have "periplasmic space" of sorts enclosed by cross-linked peptidoglycan--more leaky, however.

S-layers lie outside OM (G-) or PG (G+), and may be the only wall layer in some archaea. Role: Protection? Adherence?

Amorphous, loose layer. Can be polysaccharide (most common) or polypeptide. Prevents dehydration (general), can promote adhesion (Strep. mutans/teeth), prevent phagocytosis (Strep. pneumoniae, Neisseria meningitidis, B. anthracis, Haemophilis influenzae).

"Virulence factors"--may be conditionally produced; environmental sensing

Although glycoproteins very common in eucaryotes, rare in bacteria

2-3 types of motility: gliding motility, swimming, and "other" (e.g., marine bacteria that swim but have no apparent flagella)

Highly complex structure; requires approximately 40-50 gene products for its formation