7. Bacterial Cell Walls

• thick layer of peptidoglycan
• teichoic acids - sugar/amino polymers that give overall negative charge to the cell surface and bind Ca++ and Mg++ for eventual uptake, ~50% of mass of cell envelope
• lipoteichoic acid - covalently anchored in cytoplasmic membrane

• thin layer of peptidoglycan
• outer membrane attached to PG via lipoproteins, has porins (size dependent solute exclusion) for permeability
• outer leaflet has lipopolysaccharide (LPS)

• lipid A: fatty acids linked to glucosamine phosphate via ester amine bond, called endotoxin
• core polysaccharide: contains hexoses, heptoses and ketodeoxyoctonate
• O-specific polysaccharide (O-antigen): species-specific, made of hexoses in branched units that repeat

protects cells from bile salts, hydrophobic antibiotics and complement activation

• one large macromolecule
• glycan chains run around circumference of the cell
• peptide crosslinks connect adjacent glycan strands

• NAG+NAM in β(1,4) linked chains
• Glycan chains attached to each other by peptide crosslinks
• contains D-amino acids
• diaminopimelic acid found only in Bacteria - has two amino groups allowing peptide crosslinking

• different in different species
• gram - :between DAP and D-Ala
• gram +: between L-Lys and D-Ala or short peptide interbridge forms crosslink
• more crosslinking yields greater rigidity

• hydrophobic carrier holds PG subunits in the inner membrane until they are added to the exisiting PG scaffold
• moves PG subunits across the cytoplasmic membrane

• 1) NAM pentapeptide attached to bactoprenol to make lipid I
• - synthesized in cytoplasm and inner membrane by Mur pathway
• 2) NAG attached to lipid I to form lipid II
• - done by MurG enzyme in cytoplasmic membrane
• 3) Lipid II translocated to periplasmic side of cytoplasmic membrane, carried by bacterprenol, mediated by flipase enzymes
• * next two steps must occur outside the cell wall without the presence of ATP
• 4) transglycosylation = linking of NAG of new monomer onto NAM of existing chain by PBP1
• - bacterprenol returns to original orientation to be recycled
• 5) transpeptidation = crosslinking new PG strand into a sheet via its peptide side chain into a PG sheet by PBP1, PBP2 and PBP3

• located in cytoplasmic membrane with soluble portion facing out or entirely in periplasmic space
• high-MW PBPs (1,2,3) - perform transglycosylation or transpeptidation outside the cytoplasmic membrane
• low-MW PBPs - peptidases that remove amino acids or break the peptide crosslinks in mature PG
• lytic transcylcosylases - break the β(1,4) linkages between NAG and NAM
• functional redundancy = several enzymes perform each reaction

• if cells are in hypotonic medium when PG is disrupted they will lyse due to osmotic pressure
• if cells are in isotonic medium when PG is disrupted, the IM stays intact and protoplasts are formed
• PG sacculus retains its shape after cell lysis, but PG is not a static structure - protects against lysis

• In normal transpeptidation:
• serine residue on PBP forms covalent intermediate with a pentapeptide, released free D-ala
• the PBP D-ala intermediate is attacked by the NH2 group of DAP on a neighboring peptide to form a crosslink and release the PBP enzyme

• blocks the transpeptidation step of PG synthesis:
• mimics the D-ala-D-ala residues of PG peptides and is attacked as a substrate
• β-lactam ring is broken during attack by the PBP, forming an irreversible intermediate with the PBP

• PG is continuously synthesized and broken down
• the the presence of a β-lactam, transglycosylation proceeds normally but the glycan strands are not crosslinked, so the PG becomes weaker
• PG lysis proceeds normally, balance is shifted toward breakdown of the cell wall

mediated by β-lactamase enzymes - break the ring and inactivate the antibiotic

• PBPs help to generate characteristic cell shapes
• by knocking out one or several PBP genes at a time, investigators have found several effects on cell shape
• PBP localization conincides with predicted functions