PMID: 8939691Sep 1, 1996Paper

Mechanism of aminoglycoside 3'-phosphotransferase type IIIa: His188 is not a phosphate-accepting residue

Chemistry & Biology
P R ThompsonG D Wright

Abstract

The enzyme aminoglycoside 3'-phosphotransferase Type IIIa (APH(3')-IIIa), confers resistance to many aminoglycoside antibiotics by regiospecific phosphorylation of their hydroxyl groups. The chemical mechanism of phosphoryl transfer is unknown. Based on sequence homology, it has been suggested that a conserved His residue, His188, could be phosphorylated by ATP, and this phospho-His would transfer the phosphate to the incoming aminoglycoside. We have used chemical modification, site-directed mutagenesis and positional isotope exchange methods to probe the mechanism of phosphoryl transfer by APH(3')-IIIa. Chemical modification by diethylpyrocarbonate implicated His in aminoglycoside phosphorylation by APH(3')-IIIa. We prepared His --> Ala mutants of all four His residues in APH(3')-IIIa and found minimal effects of the mutations on the steady-state phosphorylation of several aminoglycosides. One of these mutants, His188Ala, was largely insoluble when compared to the wild-type enzyme. Positional isotope exchange experiments using gamma-[18O]-ATP did not support a double-displacement mechanism. His residues are not required for aminoglycoside phosphorylation by APH(3')-IIIa. The conserved His 188 is thus not a phosphate accepting ...Continue Reading

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Citations

Jun 1, 1997·Trends in Microbiology·J Davies, G D Wright
Oct 6, 1999·Current Opinion in Microbiology·G D Wright
Sep 9, 2010·Annual Review of Genetics·Mariya Morar, Gerard D Wright
Jan 16, 1999·Chemistry & Biology·D M DaigleG D Wright
Apr 27, 2005·The Journal of Biological Chemistry·Patrick AmstutzAndreas Plückthun
Oct 6, 1997·The Journal of Biological Chemistry·D M DaigleG D Wright
Apr 2, 1999·Antimicrobial Agents and Chemotherapy·M P Mingeot-LeclercqP M Tulkens
Aug 3, 1999·Antimicrobial Agents and Chemotherapy·K TaniguchiT Sawai
Aug 22, 2001·Drug Resistance Updates : Reviews and Commentaries in Antimicrobial and Anticancer Chemotherapy·E Azucena, S Mobashery
Jun 1, 1997·Antimicrobial Agents and Chemotherapy·T M SuterN P Cianciotto

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