Chloramphenicol acetyltransferase

Chloramphenicol acetyltransferase (or CAT) is a bacterial enzyme (EC 2.3.1.28)[1] that detoxifies the antibiotic chloramphenicol and is responsible for chloramphenicol resistance in bacteria.[2] This enzyme covalently attaches an acetyl group from acetyl-CoA to chloramphenicol, which prevents chloramphenicol from binding to ribosomes. A histidine residue, located in the C-terminal section of the enzyme, plays a central role in its catalytic mechanism.

Chloramphenicol acetyltransferase
Ribbon diagram of the chloramphenicol acetyltransferase trimer with chloramphenicol bound. From PDB: 3CLA​.
Identifiers
SymbolCAT
PfamPF00302
InterProIPR001707
PROSITEPDOC00093
SCOP23cla / SCOPe / SUPFAM
Chloramphenicol acetyltransferase
Identifiers
EC number2.3.1.28
CAS number9040-07-7
Databases
IntEnzIntEnz view
BRENDABRENDA entry
ExPASyNiceZyme view
KEGGKEGG entry
MetaCycmetabolic pathway
PRIAMprofile
PDB structuresRCSB PDB PDBe PDBsum
Gene OntologyAmiGO / QuickGO

The crystal structure of the type III enzyme from Escherichia coli with chloramphenicol bound has been determined. CAT is a trimer of identical subunits (monomer Mr 25,000) and the trimeric structure is stabilised by a number of hydrogen bonds, some of which result in the extension of a beta-sheet across the subunit interface. Chloramphenicol binds in a deep pocket located at the boundary between adjacent subunits of the trimer, such that the majority of residues forming the binding pocket belong to one subunit while the catalytically essential histidine belongs to the adjacent subunit. His195 is appropriately positioned to act as a general base catalyst in the reaction, and the required tautomeric stabilisation is provided by an unusual interaction with a main-chain carbonyl oxygen.[3]

Application

CAT is used as a reporter system to measure the level of a promoter or its tissue-specific expression. The CAT assay involves monitoring acetylation of radioactively labeled chloramphenicol on a TLC plate; CAT activity is determined by looking for the acetylated forms of chloramphenicol, which have a significantly increased migration rate as compared to the unacetylated form.[4]

References
  1. Engel J, Prockop DJ (1991). "The zipper-like folding of collagen triple helices and the effects of mutations that disrupt the zipper". Annu. Rev. Biophys. Biophys. Chem. 20 (1): 137–152. doi:10.1146/annurev.bb.20.060191.001033. PMID 1867713.
  2. Shaw WV, Packman LC, Burleigh BD, Dell A, Morris HR, Hartley BS (1979). "Primary structure of a chloramphenicol acetyltransferase specified by R plasmids". Nature. 282 (5741): 870–2. Bibcode:1979Natur.282..870S. doi:10.1038/282870a0. PMID 390404.
  3. Leslie AG (1990). "Refined crystal structure of type III chloramphenicol acetyltransferase at 1.75 A resolution". J. Mol. Biol. 213 (1): 167–186. doi:10.1016/S0022-2836(05)80129-9. PMID 2187098.
  4. Gorman, CM; Moffat LF; Howard BH (1982). "Recombinant genomes which express chloramphenicol acetyltransferase in mammalian cells". Mol. Cell. Biol. 2 (9): 1044–1051. doi:10.1128/MCB.2.9.1044. PMC 369897. PMID 6960240.
This article incorporates text from the public domain Pfam and InterPro: IPR001707
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