We demonstrate here the catalytic activity and subcellular localization of the Nm23-H4 protein, product of nm23-H4, a new member of the human nm23/nucleoside diphosphate (NDP) kinase gene family (Milon, L., Rousseau-Merck, M., Munier, A., Erent, M., Lascu, I., Capeau, J., and Lacombe, M. L. (1997) Hum. Genet. 99, 550-557). Nm3-H4 was synthesized in escherichia coli as the full-length protein and as a truncated form missing the N-terminal extension characteristic of mitochondrial targeting. The truncated form possesses NDP kinase activity, whereas the full-length protein is inactive, suggesting that the extension prevents enzyme folding and/or activity. X-ray crystallographic analysis was performed on active truncated Nm23-H4. Like other eukaryotic NDP kinases, it is a hexamer. Nm23-H4 naturally possesses a serine residue at position 129, equivalent to the K-pn mutation of the Drosophila NDP kinase. The x-ray structure shows that the presence of Ser(129) has local structural effects that weaken subunit interactions. Site-directed mutagenesis shows that the serine is responsible for the lability of Nm23-H4 to heat and urea treatment, because the S129P mutant is greatly stabilized. Examination of human embryonic kidney 293 cells transfected with green fluorescent protein fusions by confocal microscopy shows a specific mitochondrial localization of Nm23-H4 that was also demonstrated by Western blot analysis of subcellular fractions of these cells. Import into mitochondria is accompanied by cleavage of the N-terminal extension that results in NDP kinase activity. Submitochondrial fractionation indicates that Nm23-H4 is associated with mitochondrial membranes, possibly to the contact sites between the outer and inner membranes.
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==About this Structure==
==About this Structure==
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[[Category: Nm23]]
[[Category: Nm23]]
[[Category: Nucleoside diphosphate kinase]]
[[Category: Nucleoside diphosphate kinase]]
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Revision as of 21:43, 30 June 2008
This article has been automatically seeded. Changes to this page should pertain to the PDB entry only and not to the protein or biomolecule in general.
1EHW is a Single protein structure of sequence from Homo sapiens. Full crystallographic information is available from OCA.
Reference
The human nm23-H4 gene product is a mitochondrial nucleoside diphosphate kinase., Milon L, Meyer P, Chiadmi M, Munier A, Johansson M, Karlsson A, Lascu I, Capeau J, Janin J, Lacombe ML, J Biol Chem. 2000 May 12;275(19):14264-72. PMID:10799505
