Mechanistic plasticity of DNA topoisomerase IB

Phosphate electrostatics dictate the need for a catalytic arginine

Ligeng Tian, Christopher D. Claeboe, Sidney Hecht, Stewart Shuman

Research output: Contribution to journalArticle

30 Citations (Scopus)

Abstract

Four conserved amino acids of type IB topoisomerases (Arg130, Lys167, Arg223, and His265 in vaccinia topoisomerase) catalyze the attack by tyrosine on the scissile phosphodiester to form a DNA-(3′-phosphotyrosyl)-enzyme intermediate. The mechanism entails general acid catalysis (by Lys167 and Arg130) and transition-state stabilization (via contact of His265 with the pro-Sp oxygen). Here we query the function of Arg223, which accelerates transesterification by a factor of 105. The requirement for Arg223 is alleviated by a neutral Sp methylphosphonate (MeP) linkage at the cleavage site. Arg223 is not required for the 30,000-fold activation of the latent endonuclease activity of topoisomerase by the Sp MeP. The rate of autohydrolysis by the DNA-(3′-MeP)-topoisomerase intermediate approaches 10% of the rate of religation to a 5′-OH DNA strand. These findings underscore the importance of transition-state electrostatics in determining the composition of the active site and dictating the balance between strand transferase and hydrolase functions.

Original languageEnglish (US)
Pages (from-to)513-520
Number of pages8
JournalStructure
Volume13
Issue number4
DOIs
StatePublished - Apr 2005
Externally publishedYes

Fingerprint

Type I DNA Topoisomerase
Static Electricity
Arginine
Phosphates
DNA
Vaccinia
Endonucleases
Hydrolases
Transferases
Catalysis
Tyrosine
Catalytic Domain
Oxygen
Amino Acids
Acids
Enzymes
methylphosphonic acid

ASJC Scopus subject areas

  • Molecular Biology
  • Structural Biology

Cite this

Mechanistic plasticity of DNA topoisomerase IB : Phosphate electrostatics dictate the need for a catalytic arginine. / Tian, Ligeng; Claeboe, Christopher D.; Hecht, Sidney; Shuman, Stewart.

In: Structure, Vol. 13, No. 4, 04.2005, p. 513-520.

Research output: Contribution to journalArticle

Tian, Ligeng ; Claeboe, Christopher D. ; Hecht, Sidney ; Shuman, Stewart. / Mechanistic plasticity of DNA topoisomerase IB : Phosphate electrostatics dictate the need for a catalytic arginine. In: Structure. 2005 ; Vol. 13, No. 4. pp. 513-520.
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