Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
2
pubmed:dateCreated
2001-1-23
pubmed:abstractText
Phosphorylation of the nonstructural NS5A protein is highly conserved among hepatitis C virus (HCV) genotypes. However, the precise site or sites of phosphorylation of NS5A have not been determined, and the functional significance of phosphorylation remains unknown. Here, we showed by two-dimensional phosphopeptide mapping that a protein kinase or kinases present in yeast, insect, and mammalian cells phosphorylated a highly purified HCV genotype 1b NS5A from insect cells on identical serine residues. We identified a major phosphopeptide (corresponding to amino acids 2193-2212 of the HCV 1b polyprotein) by using negative-ion electrospray ionization-microcapillary high performance liquid chromatography-mass spectrometry. The elution time of the phosphopeptide determined by negative-ion electrospray ionization-mass spectrometry corresponded with the elution time of the majority of (32)P-label that was incorporated into the phosphopeptide by an in vitro kinase reaction. Subsequent analysis of the peak fraction by automated positive-ion electrospray ionization-tandem mass spectrometry revealed that Ser(2194) was the major phosphorylated residue on the phosphopeptide GpSPPSLASSSASQLSAPSLK. Substitution for Ser(2194) with Ala resulted in the concomitant disappearance of major in vivo phosphorylated peptides. Ser(2194) and surrounding amino acids are highly conserved in all HCV genotypes, suggesting NS5A phosphorylation at Ser(2194) may be an important mechanism for modulating NS5A biological functions.
pubmed:grant
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Dec
pubmed:issn
0042-6822
pubmed:author
pubmed:copyrightInfo
Copyright 2000 Academic Press.
pubmed:issnType
Print
pubmed:day
20
pubmed:volume
278
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
501-13
pubmed:dateRevised
2009-11-19
pubmed:meshHeading
pubmed-meshheading:11118372-Amino Acid Sequence, pubmed-meshheading:11118372-Animals, pubmed-meshheading:11118372-Cell Line, pubmed-meshheading:11118372-Cloning, Molecular, pubmed-meshheading:11118372-Consensus Sequence, pubmed-meshheading:11118372-Conserved Sequence, pubmed-meshheading:11118372-Hepacivirus, pubmed-meshheading:11118372-Molecular Sequence Data, pubmed-meshheading:11118372-Peptide Fragments, pubmed-meshheading:11118372-Phosphopeptides, pubmed-meshheading:11118372-Phosphorylation, pubmed-meshheading:11118372-Protein Kinases, pubmed-meshheading:11118372-Recombinant Proteins, pubmed-meshheading:11118372-Saccharomyces cerevisiae, pubmed-meshheading:11118372-Sequence Alignment, pubmed-meshheading:11118372-Sequence Homology, Amino Acid, pubmed-meshheading:11118372-Serine, pubmed-meshheading:11118372-Species Specificity, pubmed-meshheading:11118372-Spodoptera, pubmed-meshheading:11118372-Transfection, pubmed-meshheading:11118372-Viral Nonstructural Proteins
pubmed:year
2000
pubmed:articleTitle
Ser(2194) is a highly conserved major phosphorylation site of the hepatitis C virus nonstructural protein NS5A.
pubmed:affiliation
Departments of Microbiology, School of Medicine, Seattle, Washington 98195, USA. honey@u.washington.edu
pubmed:publicationType
Journal Article, Research Support, U.S. Gov't, P.H.S., Research Support, Non-U.S. Gov't