Kinetic and Catalytic Properties of Dimeric KpnI DNA Methyltransferase

Bheemanaik, Shivakumara and Chandrashekaran, Siddamadappa and Nagaraja, Valakunja and Rao, Desirazu N (2003) Kinetic and Catalytic Properties of Dimeric KpnI DNA Methyltransferase. In: Journal of Biological Chemistry, 278 (10). pp. 7863-7874.

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Abstract

KpnI DNA-(N6-adenine)-methyltransferase (KpnI MTase) is a member of a restriction-modification (R-M) system in klebsiella pneumoniae and recognizes the sequence 5_-GGTACC-3_.It modifies the recognition sequence by transferring the methyl group from Sadenosyl- L-methionine (AdoMet) to the N6 position of adenine residue. KpnI MTase occurs as a dimer in solution as shown by gel filtration and chemical crosslinking analysis. The nonlinear dependence of methylation activity on enzyme concen tration indicates that the functionally active form of the enzyme is also a dimer. Product inhibition studies with KpnI MTase showed that S-adenosyl-L-homocysteine is a competitive inhibitor with respect to AdoMet and noncompetitive inhibitor with respect to DNA. The methylated DNA showed noncompetitive inhibition with respect to both DNA and AdoMet. A reduction in the rate of methylation was observed at high concentrations of duplex DNA. The kinetic analysis where AdoMet binds first followed by DNA, supports an ordered bi bi mechanism. After methyl transfer, methylated DNA issociates followed by S-adenosyl-L-homocysteine. Isotope-partitioning analysis showed that KpnI MTase-AdoMet complex is catalytically active.

Item Type:	Journal Article
Publication:	Journal of Biological Chemistry
Publisher:	American Society for Biochemistry and Molecular Biology
Keywords:	Kinetic;Catalytic Properties;KpnI DNA Methyltransferase
Department/Centre:	Division of Biological Sciences > Microbiology & Cell Biology
Date Deposited:	17 Dec 2004
Last Modified:	19 Sep 2010 04:14
URI:	http://eprints.iisc.ac.in/id/eprint/1139

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