Explorations of peptide and oligonucleotide binding sites of tyrosyl-DNA phosphodiesterase using vanadate complexes

J Med Chem. 2004 Feb 12;47(4):829-37. doi: 10.1021/jm030487x.

Abstract

Tyrosyl-DNA phosphodiesterase (Tdp1) catalyzes the hydrolysis of a phosphodiester bond between a tyrosine residue and a DNA 3' phosphate and functions as a DNA repair enzyme that cleaves stalled topoisomerase I-DNA complexes. We previously determined a procedure to crystallize a quaternary complex containing Tdp1, vanadate, a DNA oligonucleotide, and a tyrosine-containing peptide that mimics the transition state for hydrolysis of the Tdp1 substrate. Here, the ability of vanadate to accept a variety of different ligands is exploited to produce several different quaternary complexes with a variety of oligonucleotides, and peptides or a tyrosine analogue, in efforts to explore the binding properties of the Tdp1 DNA and peptide binding clefts. Eight crystal structures of Tdp1 with vanadate, oligonucleotides, and peptides or peptide analogues were determined. These structures demonstrated that Tdp1 is able to bind substituents with limited sequence variation in the polypeptide moiety and also bind oligonucleotides with sequence variation at the 3' end. Additionally, the tyrosine analogue octopamine can replace topoisomerase I derived peptides as the apical ligand to vanadate. The versatility of this system suggests that the formation of quaternary complexes around vanadate could be adapted to become a useful method for structure-based inhibitor design and has the potential to be generally applicable to other enzymes that perform chemistry on phosphate esters.

Publication types

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

MeSH terms

  • Binding Sites
  • Crystallography, X-Ray
  • DNA Topoisomerases, Type I / chemistry
  • Ligands
  • Models, Molecular
  • Octopamine / chemistry
  • Oligonucleotides / chemistry*
  • Peptides / chemistry*
  • Phosphoric Diester Hydrolases / chemistry*
  • Structure-Activity Relationship
  • Vanadates / chemistry*

Substances

  • Ligands
  • Oligonucleotides
  • Peptides
  • Octopamine
  • Vanadates
  • Phosphoric Diester Hydrolases
  • tyrosyl-DNA phosphodiesterase
  • DNA Topoisomerases, Type I