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Molecules 2018, 23(5), 1192;

Kinetic Basis of the Bifunctionality of SsoII DNA Methyltransferase

Siberian Branch of the Russian Academy of Sciences, Institute of Chemical Biology and Fundamental Medicine, Lavrentyev Ave., 8, 630090 Novosibirsk, Russia
Belozersky Institute of Physico-Chemical Biology and Chemistry Department, Lomonosov Moscow State University, Leninskye Gory, 1, 119991 Moscow, Russia
Current Address: Swiss Institute for Experimental Cancer Research, Rue du Bugnon 21, CH-1011 Lausanne, Switzerland.
Author to whom correspondence should be addressed.
Academic Editor: Junji Iwahara
Received: 6 April 2018 / Revised: 4 May 2018 / Accepted: 8 May 2018 / Published: 16 May 2018
(This article belongs to the Special Issue Protein-DNA Interactions: From Biophysics to Genomics)
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Type II restriction–modification (RM) systems are the most widespread bacterial antiviral defence mechanisms. DNA methyltransferase SsoII (M.SsoII) from a Type II RM system SsoII regulates transcription in its own RM system in addition to the methylation function. DNA with a so-called regulatory site inhibits the M.SsoII methylation activity. Using circular permutation assay, we show that M.SsoII monomer induces DNA bending of 31° at the methylation site and 46° at the regulatory site. In the M.SsoII dimer bound to the regulatory site, both protein subunits make equal contributions to the DNA bending, and both angles are in the same plane. Fluorescence of TAMRA, 2-aminopurine, and Trp was used to monitor conformational dynamics of DNA and M.SsoII under pre-steady-state conditions by stopped-flow technique. Kinetic data indicate that M.SsoII prefers the regulatory site to the methylation site at the step of initial protein–DNA complex formation. Nevertheless, in the presence of S-adenosyl-l-methionine, the induced fit is accelerated in the M.SsoII complex with the methylation site, ensuring efficient formation of the catalytically competent complex. The presence of S-adenosyl-l-methionine and large amount of the methylation sites promote efficient DNA methylation by M.SsoII despite the inhibitory effect of the regulatory site. View Full-Text
Keywords: restriction–modification system; DNA methyltransferase; enzyme kinetics; pre-steady-state kinetics; stopped-flow assay; transcription factor restriction–modification system; DNA methyltransferase; enzyme kinetics; pre-steady-state kinetics; stopped-flow assay; transcription factor

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This is an open access article distributed under the Creative Commons Attribution License which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited (CC BY 4.0).

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Timofeyeva, N.A.; Ryazanova, A.Y.; Norkin, M.V.; Oretskaya, T.S.; Fedorova, O.S.; Kubareva, E.A. Kinetic Basis of the Bifunctionality of SsoII DNA Methyltransferase. Molecules 2018, 23, 1192.

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