Compare and contrast the Gram positive and Gram negative cell structure
Gram POSITIVE = thick, outer layer of peptidoglycan, single cellular membrane
Gram NEGATIVE = thin layer of peptidoglycan, two membranes (outer and inner)
What are the two disaccharides that polymerize to make peptidoglycan?
N-acetyl glucosamine, and N-acetyl muramic acid
In Gram (+) bacteria, how many Glycine amino acids form "bridges" between glycan strands?
What is a functional consequence of this structure?
Five - these are called "pentapeptide bridges".
This structure makes the bacterial cell wall thicker and more heavily cross-linked.
What is transpeptidation?
What amino acids are involved, and in what positions are they?
Transpeptidation is the cross-linking of peptides.
The COOH of D alanine of position 4 binds to NH2 side chain of amino acid 3 on opposing strand. Gram positives have an extra bridge of 5 glycine residues.
What enzymes catalyze transpeptidase cross-linking?
Transpeptidases and carboxypeptidases
What amino acids do transpeptidases and carboxypeptidases recognize?
They recognize D-alanine to D-alanine linkages
How are transpeptidases and carboxypeptidases inhibited by beta-lactam antibiotics (like penicillin)?
They can be competitively bound by these beta-lactam molecules because these molecules have a similar structure to the natural substrate (D-Ala to D-Ala).
This effectively stops bacterial cell wall synthesis.
What are penicillin-binding proteins (PBPs)?
PBPs are a group of proteins that are characterized by their affinity for and binding of penicillin. They are a normal constituent of many bacteria; the name simply reflects the way by which the protein was discovered!
Name four beta-lactam antibiotics
3. Cephalosporin (or Cephamycin)
The bactericidal action of the beta-lactam antibiotics requires the following four steps:
1. Association of the drug w/the bacteria.
2. Penetration through the OUTER membrane and the PERIPLASMIC space (only Gm -).
3. Interaction w/penicillin-binding proteins (PBPs) on the CYTOPLASMIC membrane.
4. Activation of an autolysin that degrades the murein in the CELL WALL.
What two strategies to bacteria use to evade beta-lactam antibiotics?
1. Produce a substance that inactivates the antibiotic (eg synthesis of beta-lactamase, which cleaves penicillin)
2. Alter the target of the drug
Name three drugs that are resistant to cleavage by beta-lactamase
How is methicillin is different from penicillin?
Methicillin is not bound and hydrolysed by penicillinase (even though it has a beta-lactam ring), meaning it can kill the bacteria, even if this enzyme is present
How does the mechanism of methicillin resistance differ from that of penicillin resistance?
Bacteria changed the target of drug interaction (done in two ways):
1. Random mutations in PBP2, encoded by the mecA gene. mecA simply encodes for an alternative PBP.
2. There is a higher frequency of transposons acquired during conjugation that jump into chromosome, conferring resistance.
If you have a bacteria that synthesizes beta-lactamase, what might be an effective treatment against it?
Use beta-lactamase inhibitors in conjunction with antibiotics.
Example: augmentin - a combined therapy with amoxicillin and clavulanic acid.
Amoxicillin is used to treat stomach ulcers and infections
What is the target recognized by vancomycin?
Describe the pathway that is inhibited as a result.
Binds to D-Ala D-Ala on MurNAc of the repeating disaccharides. Vancomycin sterically inhibits transglycosylation!
How do bacteria develop vancomycin resistance?
By creating a change in the target molecule!
Some strains acquire genes via horizontal transfer to make (D-alanyl-D-serine) or (D-alanyl-D-lactate) instead.
This type of resistance is rare.
Name two antibiotics that inhibit bacterial cell membranes.
1. Polymyxin - an antibiotic produced by bacteria (used to create a niche against other bacteria). Polymyxin aggregates on the cell membrane and the lipid tail inserts into it. Polymyxin is a narrow spectrum antibiotic for gram negatives.
2. Daptomycin - is similar to polymyxin, but for Gram (+).
Daptomycin has a cyclic amino acid structure which acts similarly to polymyxin.
Loss of the bacteria's membrane potential and integrity leads to cell death.
Daptomycin is a good alternative to vancomycin for MRSA.