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论文类型：期刊论文

发表时间：2018-01-01

发表刊物：Biochemical and biophysical research communications

收录刊物：PubMed、SCIE

卷号：501

期号：3

页面范围：674-681

ISSN号：1090-2104

关键字：HD domain containing protein; 5 ' nucleotidase; Dimerization; Nucleoside 5 '; Monophosphate

摘要：HD-domain is a conserved domain, with the signature of histidine and aspartic (HD) residues doublets. HD-domain proteins may possess nucleotidase and phosphodiesterase activities, and they play important roles in signaling and nucleotide metabolism. In yeast, HD-domain proteins with nucleotidase activity remained unexplored. Here, we biochemically and structurally characterized two HD domain proteins YGK1 (YGL101W) and YB92 (YBR242W) from Saccharomyces cerevisiae as nucleoside 5'-monophosphatases, with substrate preference for deoxyribonucleoside 5'-monophosphatase over ribonucleoside 5'-monophosphatase. By determining the crystal structure of YGK1, we unveiled that YGK1 structure resembled as the crystal structure of YfbR from E.coli. Size-exclusion chromatography and crosslinking studies suggested that YGK1 and YB92 existed in the form of a dimer, respectively, which were consistent with structural observation of YGK1. Site-directed mutagenesis demonstrated that more extensive conserved residues near the divalent metal coordinating active site were essential for YGK1 activity than previous suggested. The metal coordinating His89 and Asp90, and the neighboring conserved Glu93, Glu114 and Glu145 were individually critical for catalysis. In addition, alignments suggested that three flexible loops with hydrophobic residues might be implicated in substrate selectivity to nucleoside moiety. Together, our comparative structural and mutational studies suggested that YGK1 and YB92 functioned as 5'-nucleotidases in S.cerevisiae. Copyright © 2018 Elsevier Inc. All rights reserved.