2zcj
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(New page: 200px<br /><applet load="2zcj" size="350" color="white" frame="true" align="right" spinBox="true" caption="2zcj, resolution 2.75Å" /> '''Ternary structure of...)
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Revision as of 09:54, 23 January 2008
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Ternary structure of the Glu119Gln M.HhaI, C5-Cytosine DNA methyltransferase, with unmodified DNA and AdoHcy
Overview
The role of Glu119 in S-adenosyl-l-methionine-dependent DNA, methyltransferase M.HhaI-catalyzed DNA methylation was studied. Glu119, belongs to the highly conserved Glu/Asn/Val motif found in all DNA, C5-cytosine methyltransferases, and its importance for M.HhaI function, remains untested. We show that formation of the covalent intermediate, between Cys81 and the target cytosine requires Glu119, since conversion to, Ala, Asp or Gln lowers the rate of methyl transfer 10(2)-10(6) fold., Further, unlike the wild-type M.HhaI, these mutants are not trapped by the, substrate in which the target cytosine is replaced with the, mechanism-based inhibitor 5-fluorocytosine. The DNA binding affinity for, the Glu119Asp mutant is decreased 10(3)-fold. Thus, the ability of the, enzyme to stabilize the extrahelical cytosine is coupled directly to tight, DNA binding. The structures of the ternary protein/DNA/AdoHcy complexes, for both the Glu119Ala and Glu119Gln mutants (2.70 A and 2.75 A, respectively) show that the flipped base is positioned nearly identically, with that observed in the wild-type M.HhaI complex. A single water, molecule in the Glu119Ala structure between Ala119 and the extrahelical, cytosine N3 is lacking in the Glu119Gln and wild-type M.HhaI structures, and most likely accounts for this mutant's partial activity. Glu119 has, essential roles in activating the target cytosine for nucleophilic attack, and contributes to tight DNA binding.
About this Structure
2ZCJ is a Protein complex structure of sequences from Haemophilus parahaemolyticus with as ligand. Active as DNA (cytosine-5-)-methyltransferase, with EC number 2.1.1.37 Full crystallographic information is available from OCA.
Reference
AdoMet-dependent Methyl-transfer: Glu(119) Is Essential for DNA C5-Cytosine Methyltransferase M.HhaI., Shieh FK, Reich NO, J Mol Biol. 2007 Nov 9;373(5):1157-68. Epub 2007 Aug 19. PMID:17897676
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