Mutation of a unique aspartate residue abolishes the catalytic activity but not substrate binding of the mouse N-methylpurine-DNA glycosylase (MPG)

Rabindra Roy, Tapan Biswas, James Lee, Sankar Mitra

Research output: Contribution to journalArticle

7 Citations (Scopus)

Abstract

N-Methylpurine-DNA glycosylase (MPG) initiates base excision repair in DNA by removing a variety of alkylated purine adducts. Although Asp was identified as the active site residue in various DNA glycosylases based on the crystal structure, Glu-125 in human MPG (Glu-145 in mouse MPG) was recently proposed to be the catalytic residue. Mutational analysis for all Asp residues in a truncated, fully active MPG protein showed that only Asp- 152 (Asp-132 in the human protein), which is located near the active site, is essential for catalytic activity. However, the substrate binding was not affected in the inactive Glu-152, Asn-152, and Ala-152 mutants. Furthermore, mutation of Asp-152 did not significantly affect the intrinsic tryptophan fluorescence of the enzyme and the far UV CD spectra, although a small change in the near UV CD spectra of the mutants suggests localized conformational change in the aromatic residues. We propose that in addition to Glu-145 in mouse MPG, which functions as the activator of a water molecule for nucleophilic attack, Asp-152 plays an essential role either by donating a proton to the substrate base and, thus, facilitating its release or by stabilizing the steric configuration of the active site pocket.

Original languageEnglish (US)
Pages (from-to)4278-4282
Number of pages5
JournalJournal of Biological Chemistry
Volume275
Issue number6
DOIs
StatePublished - Feb 11 2000

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Aspartic Acid
Catalyst activity
Mutation
Substrates
Catalytic Domain
Activator Appliances
DNA Glycosylases
Tryptophan
DNA Repair
Protons
Proteins
Repair
Crystal structure
Fluorescence
DNA-3-methyladenine glycosidase II
Molecules
Water
DNA
Enzymes

ASJC Scopus subject areas

  • Biochemistry

Cite this

Mutation of a unique aspartate residue abolishes the catalytic activity but not substrate binding of the mouse N-methylpurine-DNA glycosylase (MPG). / Roy, Rabindra; Biswas, Tapan; Lee, James; Mitra, Sankar.

In: Journal of Biological Chemistry, Vol. 275, No. 6, 11.02.2000, p. 4278-4282.

Research output: Contribution to journalArticle

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