2h2u
From Proteopedia
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Crystal structure of the E130Y mutant of human soluble calcium-activated nucleotidase (SCAN) with calcium ion
Overview
Mammals express a protein homologous to soluble nucleotidases used by, blood-sucking insects to inhibit host blood clotting. These vertebrate, nucleotidases may play a role in protein glycosylation. The activity of, this enzyme family is strictly dependent on calcium, which induces a, conformational change in the secreted, soluble human nucleotidase. The, crystal structure of this human enzyme was recently solved; however, the, mechanism of calcium activation and the basis for the calcium-induced, changes remain unclear. In this study, using analytical, ultracentrifugation and chemical cross-linking, we show that calcium or, strontium induce noncovalent dimerization of the soluble human enzyme. The, location and nature of the dimer interface was elucidated using a, combination of site-directed mutagenesis and chemical cross-linking, coupled with crystallographic analyses. Replacement of Ile(170), Ser(172), and Ser(226) with cysteine residues resulted in calcium-dependent, sulfhydryl-specific intermolecular cross-linking, which was not observed, after cysteine introduction at other surface locations. Analysis of a, super-active mutant, E130Y, revealed that this mutant dimerized more, readily than the wild-type enzyme. The crystal structure of the E130Y, mutant revealed that the mutated residue is found in the dimer interface., In addition, expression of the full-length nucleotidase revealed that this, membrane-bound form can also dimerize and that these dimers are stabilized, by spontaneous oxidative cross-linking of Cys(30), located between the, single transmembrane helix and the start of the soluble sequence. Thus, calcium-mediated dimerization may also represent a mechanism for, regulation of the activity of this nucleotidase in the physiological, setting of the endoplasmic reticulum or Golgi.
About this Structure
2H2U is a Single protein structure of sequence from Homo sapiens with CA as ligand. Active as Nucleoside-diphosphatase, with EC number 3.6.1.6 Full crystallographic information is available from OCA.
Reference
Calcium-dependent dimerization of human soluble calcium activated nucleotidase: characterization of the dimer interface., Yang M, Horii K, Herr AB, Kirley TL, J Biol Chem. 2006 Sep 22;281(38):28307-17. Epub 2006 Jul 11. PMID:16835225
Page seeded by OCA on Mon Nov 12 22:25:31 2007
