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Mutational analysis of conserved residues in HhaI DNA methyltransferase

Sankpal, Umesh T and Rao, Desirazu N (2002) Mutational analysis of conserved residues in HhaI DNA methyltransferase. In: Nucleic-Acids-Research, 30 (12). pp. 2628-2638.

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HhaI DNA methyltransferase belongs to the C5-cytosine methyltransferase family, which is characterized by the presence of a set of highly conserved amino acids and motifs present in an invariant order. HhaI DNA methyltransferase has been subjected to a lot of biochemical and crystallographic studies. A number of issues, especially the role of the conserved amino acids in the methyltransferase activity, have not been addressed. Using sequence comparison and structural data, a structure-guided mutagenesis approach was undertaken, to assess the role of conserved amino acids in catalysis. Site-directed mutagenesis was performed on amino acids involved in cofactor S-adenosyl-L-methionine (AdoMet) binding (Phe18, Trp41, Asp60 and Leu100). Characterization of these mutants, by in vitro /in vivo restriction assays and DNA/AdoMet binding studies, indicated that most of the residues present in the AdoMet-binding pocket were not absolutely essential. This study implies plasticity in the recognition of cofactor by HhaI DNA methyltransferase.

Item Type: Journal Article
Publication: Nucleic-Acids-Research
Publisher: Oxford University Press
Additional Information: Copyright of this article belongs to Oxford University Press
Department/Centre: Division of Biological Sciences > Biochemistry
Date Deposited: 29 Jan 2008
Last Modified: 19 Sep 2010 04:41
URI: http://eprints.iisc.ac.in/id/eprint/12598

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