Crystal structure of O6-methylguanine-DNA methyltransferase

School of Pharmaceutical Sciences, Showa University
â—‹Masaru Tsunoda Nakamura T Kazuo

O6-methylguanine, a mutagenic, carcinogenic and toxic agent, causes base pair mismatch G:T. O6-methylguanine-DNA methyltransferase (MGMT) is a ubiquitous enzyme responsible for the repair of O6-methylguanine. The enzyme catalyzes the transfer of the methyl group from O6-methylguanine to the active site cysteine, preventing the incorrect base pairing. Here we report the crystal structures of MGMT from hyperthermophilic archaeon Sulfolobus tokodaii (StMGMT) to reveal the reaction mechanism of the methyl group transfer.
The crystal structure of StMGMT was determined by single-wavelength anomalous dispersion method at 2.0 Å resolution. In order to elucidate the reaction steps in detail, crystals of StMGMT were soaked in a solution of O6-methylguanine for four different time periods, and then they were used for X-ray diffraction experiments. The structures were determined by molecular replacement method using the apo-structure of StMGMT as a search model. Comparing the structures, we found that a hydrogen bond network ( Glu-His-Water-Cys ) in the middle of the enzyme increases the reactivity of the cysteine residue in the active site. After the reaction the network has been disrupted due to S-methylcysteine production. In the crystal structure of mutant C120S, another hydrogen bond network ( Glu-His-Water-Ser ) similar to that of the wild-type enzyme is observed in the active site, but the methytransfer reaction does not occur. It is concluded that the hydrogen bond network is utilized for the activation of cysteine residue to facilitate the methyl group transfer.