Dipole-dipole interaction stabilizes the transition state of apurinic/apyrimidinic endonuclease - Abasic site interaction

Sanjay Adhikari, Aykut Üren, Rabindra Roy

Research output: Contribution to journalArticle

11 Scopus citations

Abstract

Humanapurinic/apyrimidinic (AP) endonuclease (hAPE) initiates the repair of an abasic site (AP site). To gain insight into the mechanisms of damage recognition of hAPE, we conducted surface plasmon resonance spectroscopy to study the thermodynamics and kinetics of its interaction with substrate DNA containing an abasic site (AP DNA). The affinity of hAPE binding toward DNA increased as much as 6-fold after replacing a single adenine (equilibrium dissociation constant, KD, 5.3 nM) with an AP site (KD, 0.87 nM). The enzyme-substrate complex formation appears to be thermodynamically stabilized and favored by a large change in Gibbs free energy, ΔG° (-50 kJ/mol). The latter is supported by a high negative change in enthalpy, ΔH° (-43 kJ/mol) and also positive change in entropy, ΔS° (24 J/(K mol)), and thus the binding process is spontaneous at all temperatures. Analysis of kinetic parameters reveals small enthalpy of activation for association, ΔH°Dagger; ass (-17 kJ/mol), and activation energy for association (Ea, -14 kJ/mol) when compared with the enthalpy of activation for dissociation, ΔH° Dagger; diss (26 kJ/mol), and activation energy in the reverse direction (Ed, 28 kJ/mol). Furthermore, varying concentration of KCl showed an increase in binding affinity at low concentration but complete abrogation of the binding at higher concentration, implying the importance of hydrophobic, but predominately ionic, forces in the Michaelis-Menten complex formation. Thus, low activation energy and the enthalpy of activation, which are perhaps a result of dipole-dipole interactions, play critical roles in AP site binding of APE.

Original languageEnglish (US)
Pages (from-to)1334-1339
Number of pages6
JournalJournal of Biological Chemistry
Volume283
Issue number3
DOIs
StatePublished - Jan 18 2008

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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