title: In vitro characterization of the HSV-1 UL53 gene product.
abstract: The UL53 gene is the locus altered in many syncytial mutants of herpes simplex virus type 1 (HSV-1). However, the protein encoded by this gene has not been characterized. In this study, the UL53 protein was produced by in vitro translation of in vitro-transcribed UL53 RNA. Post-translational processing of the protein was studied by translation in the presence of pancreatic microsomal membranes. These microsomes carry out the processing steps that normally occur in the rough endoplasmic reticulum. The unprocessed protein had an apparent molecular weight of 27K, whereas the microsomally processed form had an apparent molecular weight of 36K. Two types of post-translational modification were detected: Addition of N-linked oligosaccharides and cleavage of an N-terminal signal sequence. N-linked glycosylation occurred in the first 112 residues of the protein, consistent with the presence of N-linked glycosylation signals at residues 48 and 58. Signal sequence cleavage occurred after residue 30. A membrane-binding, possibly transmembrane, domain was found between residues 113 and 170, probably consisting of the hydrophobic sequence 125-139. These results establish that the N-terminal domain of the UL53 protein, which is the site of those syncytial mutations that have been sequenced, is on the interior side of the microsomal membranes, which is topologically equivalent to the lumen of the rough endoplasmic reticulum and to the extracellular side of the plasma membrane. Additional hydrophobic, possibly transmembrane, domains exist nearer the C-terminus of the protein. It also was found that the in vitro-translated UL53 protein aggregated when heated, even in the presence of SDS. This property was mapped to the C-terminal one-third of the protein.
authors.affiliation: Department of Immunology and Microbiology, Wayne State University Medical School, Detroit, Michigan 48201.
authors.forename: R
authors.initials: R
authors.lastname: Ramaswamy
authors.forename: T C
authors.initials: TC
authors.lastname: Holland
mesh_terms: D000595:Amino Acid Sequence; D002254:Carbonates / Q000494:pharmacology; D003001:Cloning, Molecular; D004591:Electrophoresis, Polyacrylamide Gel; D006026:Glycoside Hydrolases / Q000378:metabolism; D008969:Molecular Sequence Data; D014176:Protein Biosynthesis; D015183:Restriction Mapping; D018139:Simplexvirus / Q000235:genetics*; D014764:Viral Proteins / Q000737:chemistry / Q000235:genetics*
chemical_list: D002254:Carbonates; C072448:UL53 protein, Human herpesvirus 1; D014764:Viral Proteins; C005686:sodium carbonate; D006026:Glycoside Hydrolases