The chemokine receptor homologue encoded by US27 of human cytomegalovirus is heavily glycosylated and is present in infected human foreskin fibroblasts and enveloped virus particles.
Human cytomegalovirus (HCMV), a member of the beta-herpesvirus family, encodes four homologues of cellular G protein-coupled receptors (GPCRs). One of these, the protein product of HCMV open reading frame (ORF) UL33, has been identified in HCMV-infected cells and virus particles and shown to be heat-aggregatable and N-glycosylated. Another, the product of ORF US28, has been functionally characterized as a beta-chemokine receptor. Here we report the use of RT-PCR, coupled in vitro transcription-translation, immunoprecipitation, and Western immunoassays to (i) show that RNA from the open reading frame US27 appears predominantly during the late phase of replication; (ii) identify the protein encoded by HCMV US27 in infected cells and enveloped virus particles; (iii) demonstrate that the US27-encoded protein is heterogeneously N-glycosylated and resolves as two species following treatment with peptide N-glycosidase F; and (iv) show that both the recombinant and deglycoylated infected cell US27 protein aggregate when heated in the presence of SDS prior to electrophoresis in polyacrylamide gels, a property which is abrogated with the addition of urea to sample buffer.
Margulies
Barry J
BJ
Towson University Herpes Virus Lab, Department of Biological Sciences, Towson University, Towson, MD 21252, USA. bjmarg@alum.mit.edu
Gibson
Wade
W
D002478:Cells, Cultured; D003587:Cytomegalovirus / Q000472:pathogenicity / Q000502:physiology*; D003586:Cytomegalovirus Infections / Q000821:virology*; D005347:Fibroblasts / Q000378:metabolism / Q000821:virology; D006031:Glycosylation; D006801:Humans; D019707:Receptors, Chemokine / Q000378:metabolism*; D014764:Viral Proteins / Q000378:metabolism*; D014774:Virulence; D014779:Virus Replication
D019707:Receptors, Chemokine; C461644:US27 protein, human cytomegalovirus; D014764:Viral Proteins