Question

Ways in which Chemical Control Agents May Affect Bacteria

1. Describe one way an antibiotic can inhibit peptidoglycan synthesis, state how that ultimately kills the bacterium, and give an example of such an antibiotic.

2. Describe one way an antibiotic can alter bacterial ribosomes, state how that ultimately inhibits or kills the bacterium, and give an example of such an antibiotic.

3. Describe one way an antibiotic can interfere with bacterial DNA synthesis, state how that ultimately kills the bacterium, and give an example of such an antibiotic.

Ways in which Bacteria May Resist Chemical Control Agent

1. Briefly describe 3 different mechanisms, as a result of mutation or horizontal gene transfer in a bacterium, that may enable that bacterium to resist an antibiotic

Answer 1

A. Chemical Control Agents that affect bacteria

1. Peptidoglycan, a cell wall component is synthesized by crosslinking of its glycan strand. Transglycosidases catalyze this reaction leading to formation of polymers, which will then cross link to each other. Penicillin binding proteins are required for the cross linking of D-alanyl-alanine to glycine residue in peptidoglycan synthesis as they bind D-alany-D-alanine residues. Beta lactams are antibiotics that inhibit penicillin binding proteins. The beta lactam ring present in these antibiotics has a structure similar to D-alanyl-D-alanine. Hence, this beta lactam ring will interact with penicillin binding proteins. This prevents the interaction between D-alanyl-D-alanine with glycine. Further, penicillin binding proteins are sequestered and not available for peptidoglycan synthesis, thereby inhibiting peptidoglycan synthesis. Thus, there is no synthesis of cell wall leading to death of the bacterium. Beta lactam antibiotics include penicillin and cephalosporin.

2. Bacterial ribosomes (70S) are made up of two components- a smaller subunit of 30S and large subunit of 50S. Antibiotics can have an effect on any of these components, which then inhibits protein synthesis. Ribosomes are the sites of protein translation, which interact both with mRNA and tRNA. Macrolides are antibiotics that affect 50S subunit of ribosomes. They affect translocation step of protein synthesis. Macrolides inhibit the P (peptidyl) site on 50S ribosome, thereby inhibiting peptidyl transfer reaction and consequently inhibit translocation. This causes detachment of peptide chain prematurely which inhibits protein synthesisin the bacteria. Host cells have the 80S ribosomes that are not inhibited. Example of macrolides is clathromycin, and Azithromycin.

3. Quinolones are inhibitors of DNA replication which is required to synthesize DNA. One of the quinolones is the fluoroquinolones (FQ). FQ inhibits the DNA gyrase enzyme involved in DNA replication. DNA gyrase subunit B introduces negative supercoils in DNA after double stranded nicks. The nicked ends are then resealed by subunit A of DNA gyrase. FQ inhibits DNA gyrase by binding to DNA gyrase subunit and thereby inhibits DNA replication. Ciprofloxacin and levofloxacin are examples of FQ antibiotics. Host cells do not have DNA gyrase enzyme.

B: Antibiotic resistance via HGT and Mutation:

1. There are different ways in which antibiotic resistance can occur due to horizontal gene transfer or mutation.

a. Mutations can occur that alter the target site of the antibiotic. Mutations in the sequences of penicillin binding proteins involved in synthesis of peptidoglycan causes reduced interactions with beta lactam ring. This will prevent action of beta lactam antibiotics that binds to these proteins. Amino glycosides bind to 30S subunit. If the binding site of the antibiotic on the 30S ribosome is mutated, the antibiotic can no longer bind to it, thereby inhibiting its action on bacteria.

b. The bacteria can obtain ability to synthesize enzymes that inactivate the antibiotic as a result of horizontal gene transfer. Horizontal gene transfer is transfer of gene between microorganism via conjugation, transduction and transformation. Penicillinase is an enzyme that can cleave beta lactam ring of penicillin, a beta lactam antibiotic. Cephalosporinase is an enzyme that inactivates cephalosporin antibiotic. Chloramphenicol transacetylase is an enzyme that inactivates chloramphenicol by acetylating its hydroxyl groups. As a result, chloramphenicol will no longer be able to inhibit 50S ribosomal subunit.

c. Bacteria can also synthesize new efflux pumps for antibiotics as a result of HGT. As a result, they are able to efflux out the antibiotics from the cell. Hence even though antibiotics enter the cell, these efflux pumps can remove them easily from the cell, thereby maintaining very low concentration of these antibiotics intracellularly. This low concentration will not be able to inhibit cellular bacterial growth. Resistance can be gained to macrolides, tetracyclines, and FQ by synthesis of these efflux pumps.