Crosslinking of (Cytosine-5)-DNA Methyltransferase SsoII and its Complexes with Specific DNA Duplexes Provides an Insight into Their Structures

Abstract

(Cytosine-5)-DNA methyltransferase SsoII (M.SsoII) functions as a methyltransferase and also as a transcription factor. Chemical and photochemical crosslinking was used for exploring the structure of M.SsoII–DNA complexes and M.SsoII in the absence of DNA. Photocrosslinking with 4-(N-maleimido)benzophenone demonstrated that in the M.SsoII complex with DNA containing the regulatory site, the M.SsoII region responsible for methylation was bound to DNA flanking the regulatory site, which contained no methylation sequence. This required high flexibility of the linker connecting the M.SsoII N-terminal domain and the M.SsoII region responsible for methylation. The flexibility was demonstrated by crosslinking with bis-maleimidoethane and 1,11-bis-maleimidotetraethyleneglycol.

Keywords:

• DNA methyltransferase SsoII
• protein–protein crosslinking
• protein–DNA crosslinking
• protein–DNA complexes

Acknowledgments

This work has been supported by the German Research Foundation (DFG) and the Russian Foundation for Basic Research (RFBR) program “International Research Training Groups” (grants GRK 1384, 11-04-91336), Russian Foundation for Basic Research (grant 10-04-01578), and grant for Leading Scientific Schools (3314.2010.4). The authors are grateful to Dr. Karyagina and Dr. Solonin for the kind gift to M.SsoII and M.Ecl18kI plasmids.