Discrimination of lesion removal of N-methylpurine-DNA glycosylase revealed by a potent neutralizing monoclonal antibody

Sanjay Adhikari, Stephen J. Kennel, Gargi Roy, Partha S. Mitra, Sankar Mitra, Rabindra Roy

Research output: Contribution to journalArticlepeer-review

8 Scopus citations


N-Methylpurine-DNA glycosylase (MPG), a ubiquitous DNA repair enzyme, initiates excision repair of several N-alkylpurine adducts, induced by alkylating chemotherapeutics, and deaminated and lipid peroxidation-induced purine adducts. We have generated monoclonal antibodies (moAbs) against human MPG. Twelve independent hybridoma clones were characterized, which, except 520-16A, are identical based on epitope exclusion assay. Four moAbs, including 520-2A, 520-3A, 520-16A, and 520-26A, have high affinity (KD ∼ 0.3-1.6 nM), and their subtypes were IgG2a, IgG1, IgG2a, and IgG2b, respectively. moAb 520-3A recognizes the sequence 52AQAPCPRERCLGPP66T, an epitope exclusively present in the N-terminal extension of human MPG. We found that moAb 520-3A significantly inhibited MPG's enzymatic activity towards different substrates, such as hypoxanthine, 1,N6ethenoadenine and methylated bases, which represent different classes of DNA damage, however, with different efficiencies. Real-time binding experiments using surface plasmon resonance (SPR) spectroscopy showed that the pronounced inhibition of activity was not in the substrate-binding step. Single turnover kinetics (STO) revealed that the inhibition was at the catalytic step. Since we found that this antibody has an epitope in the N-terminal tail, the latter appears to have an important role in substrate discrimination, however, with a differential effect on different substrates.

Original languageEnglish (US)
Pages (from-to)31-39
Number of pages9
JournalDNA Repair
Issue number1
StatePublished - Jan 1 2008


  • Base excision repair
  • Epitope mapping
  • Inhibition
  • MPG
  • Substrate specificity

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology


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