Absence of a phosphorylated intermediate during ATP hydrolysis by Escherichia coli transcription termination protein rho

J Biol Chem. 1986 Dec 5;261(34):15906-9.

Abstract

We have established the suitability of adenosine 5'-O-(gamma-thio)triphosphate(ATP gamma S) as an analog of ATP for the nucleoside triphosphatase activity of Escherichia coli transcription termination protein rho (EC 3.6.1.3). Steady-state analysis gives a Vmax of 1.5 mumol min-1 mg-1, 9% of the value with MgATP as substrate, and indicates that ATP gamma S binds as tightly (based on Km and Ki versus ATP) to rho as does ATP. (gamma-S)[beta gamma-17O,gamma-17O,gamma-18O]ATP gamma S was used as substrate to produce chiral product inorganic [17O,18O]thiophosphate and determine the stereochemical course of the hydrolysis. The results of this determination, inversion at the thiophosphoryl phosphorus, indicate that the enzymatic hydrolysis of ATP by rho consists of a direct transfer of the phospho group to water without the existence of a phosphoenzyme or phospho-RNA intermediate.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, Non-P.H.S.
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Adenosine Triphosphate / analogs & derivatives
  • Adenosine Triphosphate / metabolism*
  • Escherichia coli / metabolism
  • Hydrolysis
  • Kinetics
  • Molecular Conformation
  • Phosphorylation
  • Rho Factor / pharmacology*
  • Transcription Factors / pharmacology*

Substances

  • Rho Factor
  • Transcription Factors
  • adenosine 5'-O-(3-thiotriphosphate)
  • Adenosine Triphosphate