2c7p: Difference between revisions

DNA base flipping is an important mechanism in molecular enzymology, but, its study is limited by the lack of an accessible and reliable diagnostic, technique. A series of crystalline complexes of a DNA methyltransferase, M.HhaI, and its cognate DNA, in which a fluorescent nucleobase analogue, 2-aminopurine (AP), occupies defined positions with respect the target, flipped base, have been prepared and their structures determined at higher, than 2 A resolution. From time-resolved fluorescence measurements of these, single crystals, we have established that the fluorescence decay function, of AP shows a pronounced, characteristic response to base flipping: the, loss of the very short (approximately 100 ps) decay component and the, large increase in the amplitude of the long (approximately 10 ns), component. When AP is positioned at sites other than the target site, this, response is not seen. Most significantly, we have shown that the same, clear response is apparent when M.HhaI complexes with DNA in solution, giving an unambiguous signal of base flipping. Analysis of the AP, fluorescence decay function reveals conformational heterogeneity in the, DNA-enzyme complexes that cannot be discerned from the present X-ray, structures.

2C7P is a [http://en.wikipedia.org/wiki/Protein_complex Protein complex] structure of sequences from [http://en.wikipedia.org/wiki/Haemophilus_haemolyticus Haemophilus haemolyticus] with SO4, SAH, EPE and CIT as [http://en.wikipedia.org/wiki/ligands ligands]. Active as [http://en.wikipedia.org/wiki/DNA_(cytosine-5-)-methyltransferase DNA (cytosine-5-)-methyltransferase], with EC number [http://www.brenda-enzymes.info/php/result_flat.php4?ecno=2.1.1.37 2.1.1.37] Structure known Active Site: AC1. Full crystallographic information is available from [http://ispc.weizmann.ac.il/oca-bin/ocashort?id=2C7P OCA].  

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