1887

Abstract

The gene product of the human immunodeficiency virus (HIV) is suggested to be a negative factor involved in down-regulating viral expression by a mechanism in which the correct conformation of the protein is essential. The protein expressed by vaccinia virus recombinants is phosphorylated by protein kinase C. We investigated the synthesis of the protein and its state of phosphorylation during HIV-1 infection of a T4 cell line (CEM cells). Maximum synthesis of viral proteins occurred 3 days after infection, when more than 90% of cells were producing viral proteins. The synthesis of the protein was detected in parallel with the and proteins. As expected, the protein was myristylated but not phosphorylated, and its half-life was less than 1 h. By the use of the polymerase chain reaction technique, we isolated and sequenced the gene of this HIV-1 stock. Two significant mutations were observed. Firstly, threonine, at amino acid number 15, the site of phosphorylation by protein kinase C, was mutated into an alanine, and secondly aspartic acid of the tetrapeptide WRFD, which is probably involved in GTP binding, was mutated into an asparagine. The mutated gene was expressed in a vaccinia virus system, in which it was not phosphorylated and its half-life was dramatically reduced compared to the wild-type gene product. Furthermore, down-regulation of CD4 cell surface expression was no longer affected by the mutated gene. These results emphasize that phosphorylation of the protein provides an efficient test to monitor its biological activity.

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/content/journal/jgv/10.1099/0022-1317-71-10-2273
1990-10-01
2019-11-21
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http://instance.metastore.ingenta.com/content/journal/jgv/10.1099/0022-1317-71-10-2273
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