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Analyst. 2011 Mar 21;136(6):1227-33. PubMed; doi 10.1039/c0an00545b

The study of the interaction of (cytosine-5)-DNA methyltransferase SsoII with DNA by acoustic method.

Ryazanova, A. Y.; Kubareva, E. A.; Grman, I.; Lavrova, N. V.; Ryazanova, E. M.; Oretskaya, T. S.; Hianik, T.

The interaction of (cytosine-5)-DNA methyltransferase SsoII (M.SsoII) with double-stranded DNA was studied by means of thickness shear mode acoustic method (TSM) and gel electrophoresis. M.SsoII recognizes in double-stranded DNA the methylation site 5'-CCNGG-3' (N=A, C, G, T) and methylates the inner cytosine residue. M.SsoII also acts as a transcription factor via binding to the regulatory site 5'-AGGACAAATTGTCCT-3' in the promoter region of SsoII restriction-modification system. We designed three 60-mer biotinylated DNA duplexes: with the methylation site (60met), with the regulatory site (60reg), and without a specific binding site (60oct). A strong binding of M.SsoII with each one of the studied DNA immobilized on the TSM transducer has been shown. The equilibrium dissociation constants, K(D), of the M.SsoII-DNA complexes decreased in the order 60oct>60reg>60met, suggesting a higher stability of M.SsoII-60met complex in comparison with the others. The association rate constant, k(a), was also higher for 60met, while similar values were obtained for 60reg and 60oct. The difference in the kinetic parameters for 60met and 60reg suggested a possible way of coordination between the two M.SsoII functions in a cell.



 


  Московский Государственный Университет имени М.В.Ломоносова



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© 2011 Факультет биоинженерии и биоинформатики
Московского Государственного Университета имени М.В.Ломоносова


 





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