JVI Figure table search 04
Home Help [Feedback] [For Subscribers] [Archive] [Search] [Contents]
This Article
Right arrow Full Text
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrowReprints and Permissions
Right arrow Copyright Information
Right arrow Books from ASM Press
Right arrow MicrobeWorld
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Taylor, D. R.
Right arrow Articles by Mathews, M. B.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Taylor, D. R.
Right arrow Articles by Mathews, M. B.

 Previous Article  |  Next Article 

Journal of Virology, February 2001, p. 1265-1273, Vol. 75, No. 3
0022-538X/01/$04.00+0   DOI: 10.1128/JVI.75.3.1265-1273.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.

Hepatitis C Virus Envelope Protein E2 Does Not Inhibit PKR by Simple Competition with Autophosphorylation Sites in the RNA-Binding Domain

Deborah R. Taylor,1,2,3 Bin Tian,4 Patrick R. Romano,5,dagger Alan G. Hinnebusch,5 Michael M. C. Lai,3 and Michael B. Mathews1,4,*

Cold Spring Harbor Laboratory, Cold Spring Harbor, New York 117241; Genetics Program, State University of New York at Stony Brook, Stony Brook, New York 117942; Department of Molecular Microbiology and Immunology, Howard Hughes Medical Institute, University of Southern California School of Medicine, University of Southern California, Los Angeles, California 900333; Department of Biochemistry and Molecular Biology, New Jersey Medical School, University of Medicine and Dentistry of New Jersey, Newark, New Jersey 071034; and Laboratory of Eukaryotic Gene Regulation, National Institute of Child Health and Human Development, Bethesda, Maryland 208925

Received 15 August 2000/Accepted 31 October 2000

Double-stranded-RNA (dsRNA)-dependent protein kinase PKR is induced by interferon and activated upon autophosphorylation. We previously identified four autophosphorylated amino acids and elucidated their participation in PKR activation. Three of these sites are in the central region of the protein, and one is in the kinase domain. Here we describe the identification of four additional autophosphorylated amino acids in the spacer region that separates the two dsRNA-binding motifs in the RNA-binding domain. Eight amino acids, including these autophosphorylation sites, are duplicated in hepatitis C virus (HCV) envelope protein E2. This region of E2 is required for its inhibition of PKR although the mechanism of inhibition is not known. Replacement of all four of these residues in PKR with alanines did not dramatically affect kinase activity in vitro or in yeast Saccharomyces cerevisiae. However, when coupled with mutations of serine 242 and threonines 255 and 258 in the central region, these mutations increased PKR protein expression in mammalian cells, consistent with diminished kinase activity. A synthetic peptide corresponding to this region of PKR was phosphorylated in vitro by PKR, but phosphorylation was strongly inhibited after PKR was preincubated with HCV E2. Another synthetic peptide, corresponding to the central region of PKR and containing serine 242, was also phosphorylated by active PKR, but E2 did not inhibit this peptide as efficiently. Neither of the PKR peptides was able to disrupt the HCV E2-PKR interaction. Taken together, these results show that PKR is autophosphorylated on serine 83 and threonines 88, 89, and 90, that this autophosphorylation may enhance kinase activation, and that the inhibition of PKR by HCV E2 is not solely due to duplication of and competition with these autophosphorylation sites.

* Corresponding author. Mailing address: Department of Biochemistry and Molecular Biology, New Jersey Medical School, University of Medicine and Dentistry of New Jersey, Newark, NJ 07103. Phone: (973) 972-4411. Fax: (973) 972-5594. E-mail: mathews{at}UMDNJ.edu.

dagger Present address: Jefferson Center for Biomedical Research, Doylestown, PA 18901-2697.

Journal of Virology, February 2001, p. 1265-1273, Vol. 75, No. 3
0022-538X/01/$04.00+0   DOI: 10.1128/JVI.75.3.1265-1273.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.


This article has been cited by other articles:



Home Help [Feedback] [For Subscribers] [Archive] [Search] [Contents]
J. Bacteriol. Mol. Cell. Biol. Microbiol. Mol. Biol. Rev.
Clin. Vaccine Immunol. ALL ASM JOURNALS

Copyright © 2001 by the American Society for Microbiology. All rights reserved.