Linked Life Data

14967486

Source:http://linkedlifedata.com/resource/pubmed/id/14967486

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
1
pubmed:dateCreated
2004-2-17
pubmed:abstractText
An expression system for analysis of the synthesis and processing of the E2 glycoprotein of a hepatitis C virus (HCV) genotype 1a strain was developed in transiently transfected cells. E2 proteins representing the entire length of the protein, including the transmembrane segment (E2) as well as two truncated versions (E2(660) and E2(715)), were characterized for acquisition of N-linked glycans and transport to the media of transfected cells. To investigate the utilization of the 10 potential N-linked glycosylation sites on this E2 protein, a series of mutations consisting of single or multiple (two, three, four or eight) ablations of asparagine residues in the background of the E2(660) construct were analyzed. E2(660) proteins harboring single or multiple site mutations were produced at levels similar to that of wild-type protein, but secretion of the single mutants was mildly diminished, and elimination of two or more sites dramatically reduced delivery of the protein to the media. Similar results were obtained in Huh-7 cells with respect to intracellular synthesis and secretion of the mutant proteins. Analysis of oligosaccharide composition using endoglycosidase digestion revealed that all of the glycan residues on the intracellular forms of E2(660), E2(715), and E2 contained N-linked glycans modified into high-mannose carbohydrates, in contrast to the secreted forms, which were endo H resistant. The parental E2(660) protein could be readily detected in Huh-7 cells using anti-polyhistidine or antibody to recombinant E2. In contrast, E2(660) lacking the eight N-linked glycans was expressed but not detectable with anti-E2 antibody, and proteins lacking four glycans exhibited reduced reactivity. These experiments provide direct evidence that the presence of multiple N-linked glycans is required for the proper folding of the E2 protein in the ER and secretory pathway as well as for formation of its antigenic structure.
pubmed:grant
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Feb
pubmed:issn
0042-6822
pubmed:author
pubmed:issnType
Print
pubmed:day
5
pubmed:volume
319
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
36-48
pubmed:dateRevised
2007-11-14
pubmed:meshHeading
pubmed:year
2004
pubmed:articleTitle
HCV E2 glycoprotein: mutagenesis of N-linked glycosylation sites and its effects on E2 expression and processing.
pubmed:affiliation
Department of Molecular Microbiology and Immunology, St. Louis University School of Medicine, St. Louis, MO 63104, USA. chambetj@slu.edu
pubmed:publicationType
Journal Article, Research Support, U.S. Gov't, P.H.S., Research Support, Non-U.S. Gov't