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Probing the role of correlated motions in the catalytic activity of M. HhaI by NMR
Author(s) -
Shatz Whitney,
Purdy Matt
Publication year - 2006
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.20.4.a51-b
Subject(s) - methyltransferase , dna , chemistry , methylation , dna methyltransferase , enzyme , cofactor , dna methylation , gene , biophysics , biochemistry , nuclear magnetic resonance , gene expression , biology , physics
Enzymatic DNA methylation is involved in the regulation of gene expression. Mammalian DNA methyltransferases have been identified as targets for anti‐cancer drugs. To further understand the mechanism of DNA methyltransferases, the correlation between active site and non‐active site motions of M. HhaI, a bacterial homolog, is being studied. A TROSY solution state NMR spectrum has been obtained with 15 N labeled protein, but due to its 37 kDa size the signal to noise ratio is low. Higher concentrations are needed to increase the signal strength and sharpen the broad peaks. At such high concentrations, the wild type protein is weakly soluble and dimerizes. Different combinations of DNA and cofactor, varying buffer composition and the engineering of new constructs are being explored to overcome these issues in order to obtain a triply labeled Trosy spectrum necessary for making residue assignments.