The mechanism of antibiotics

Biol 1220 Synthetic Biologyabe pressman & minoo ramanathan

the basics

• Used to kill or inhibit the growth of bacteria• Classified as bactericidal or bacteriostatic

Kill bacteria directly Prevent cell division• Classified by target specificity: Narrow-spectrum vs Broad range• Most modified chemically from original

compounds found in nature, some isolated and produced from living organisms

sites of antiobiotic action

ampicillin

• Belongs to β-lactam group of antibiotics – contain β-lactam ring

• Broad-spectrum• Penicillin derivative that inhibits bacterial cell

wall synthesis (peptidoglycan cross-linking) • Inactivates transpeptidases on the inner surface

of the bacterial cell membrane• Bactericidal only to growing E. Coli• Widespread use leads to bacterial resistance.

HOW?

ampicillin resistance

• Cleavage of β-lactam ring by β-lactamase enzyme

ampicillin resistance

• β-lactamase is encoded by the plasmid-linked bla (TEM-1) gene

• Hydrolyzes ampicillin • Ampicillin levels in culture continually

depleted

use in synthetic biology• To confirm uptake of gene (eg. of plasmids) by

bacteria• Bacterial Transformation: DNA integrates into

bacteria’s chromosome and made chemically competent

• Exogenous DNA tagged with an antibiotic resistance gene eg. β-lactamase

• Grown in medium containing ampicillin• Ampicillin resistance indicates successful

bacterial transformation

Kanamycin

• Targets 30s ribosomal subunit, causing a frameshift in every translation

• Bacteriostatic: bacterium is unable to produce any proteins correctly, leading to a halt in growth and eventually cell death

kanamycin use/resistance

• Over-use of kanamycin has led to many wild bacteria possessing resistance plasmids

• As a result of this (as well as a lot of side effects in humans), kanamycin is widely used for genetic purposes rather than medicinal purposes, especially in transgenic plants

• Resistance is often to a family of related antibiotics, and can include antibiotic-degrading enzymes or proteins protecting the 30s subunit

chloramphenicol• Bacteriostatic: functions by halting bacterial growth,

which is done by inhibiting the enzyme peptidyl transferase, a protein that assists in the binding of tRNA to the 50s ribosomal subunit

• Three methods of resistance: reduced membrane permeability, mutation of the 50s subunit, and an enzyme called chloramphenicol acetyltransferase, which inactivates chloramphenicol by covaltly linking groups

• Easy/cheap to manufacture, but unused in western countries because of possible aplastic anemia as a side effect

Sources

• http://en.wikipedia.org/wiki/Antibiotic

• http://en.wikipedia.org/wiki/Ampicillin

• http://en.wikipedia.org/wiki/Beta-lactamase

• http://www.sigmaaldrich.com/catalog/ProductDetail.do?N4=A1593|

SIAL&N5=SEARCH_CONCAT_PNO|BRAND_KEY&F=SPEC

• http://abe.leeward.hawaii.edu/Protocols/QiagenSpinprepProtocol.htm

• http://www.openwetware.org/wiki/Brown_BIOL1220:Notebook/

SynBio_in_Theory_and_Practice/Bacterial_Basics