Details

Autor(en) / Beteiligte

• McCann, Joe A. B
• Berti, Paul J

Titel

Transition-State Analysis of the DNA Repair Enzyme MutY

Ist Teil von

• Journal of the American Chemical Society, 2008-04, Vol.130 (17), p.5789-5797

Ort / Verlag

United States: American Chemical Society

Erscheinungsjahr

2008

Quelle

MEDLINE

Beschreibungen/Notizen

• The transition state (TS) structure of MutY-catalyzed DNA hydrolysis was solved using multiple kinetic isotope effect (KIE) measurements. MutY is a base excision repair enzyme which cleaves adenine from 8-oxo-G:A mismatches in vivo, and also from G:A mismatches in vitro. TS analysis of G:A-DNA hydrolysis revealed a stepwise SN1 (DN*AN ‡) mechanism proceeding through a highly reactive oxacarbenium ion intermediate which would have a lifetime in solution of <10−10 s. C−N bond cleavage is reversible; the N-glycoside bond breaks and reforms repeatedly before irreversible water attack on the oxacarbenium ion. KIEs demonstrated that MutY uses general acid catalysis by protonating N7. It enforces a 3′-exo sugar ring conformation and other sugar ring distortions to stabilize the oxacarbenium ion. Combining the experimental TS structure with the previously reported crystal structure of an abortive Michaelis complex elucidates the step-by-step catalytic sequence.