Mechanism of cis-prenyltransferase reaction probed by substrate analogues

Biochem Biophys Res Commun. 2010 Oct 1;400(4):758-62. doi: 10.1016/j.bbrc.2010.09.001. Epub 2010 Sep 7.

Abstract

Undecaprenyl pyrophosphate synthase (UPPS) is a cis-type prenyltransferases which catalyzes condensation reactions of farnesyl diphosphate (FPP) with eight isopentenyl pyrophosphate (IPP) units to generate C(55) product. In this study, we used two analogues of FPP, 2-fluoro-FPP and [1,1-(2)H(2)]FPP, to probe the reaction mechanism of Escherichia coli UPPS. The reaction rate of 2-fluoro-FPP with IPP under single-turnover condition is similar to that of FPP, consistent with the mechanism without forming a farnesyl carbocation intermediate. Moreover, the deuterium secondary KIE of 0.985±0.022 measured for UPPS reaction using [1,1-(2)H(2)]FPP supports the associative transition state. Unlike the sequential mechanism used by trans-prenyltransferases, our data demonstrate E. coli UPPS utilizes the concerted mechanism.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Alkyl and Aryl Transferases / metabolism*
  • Catalysis
  • Escherichia coli / enzymology*
  • Hemiterpenes / metabolism*
  • Organophosphorus Compounds / metabolism*
  • Polyisoprenyl Phosphates / metabolism*
  • Sesquiterpenes / metabolism*
  • Substrate Specificity
  • Transferases / metabolism*

Substances

  • 2-fluorofarnesyl diphosphate
  • Hemiterpenes
  • Organophosphorus Compounds
  • Polyisoprenyl Phosphates
  • Sesquiterpenes
  • isopentenyl pyrophosphate
  • farnesyl pyrophosphate
  • Transferases
  • Alkyl and Aryl Transferases
  • cis-prenyl transferase
  • undecaprenyl pyrophosphate synthetase