Incorporation of 12-methoxydodecanoate into the human immunodeficiency virus 1 gag polyprotein precursor inhibits its proteolytic processing and virus production in a chronically infected human lymphoid cell line.
Covalent linkage of myristate (tetradecanoate; 14:0) to the NH2-terminal glycine residue of the human immunodeficiency virus 1 (HIV-1) 55-kDa gag polyprotein precursor (Pr55gag) is necessary for its proteolytic processing and viral assembly. We have shown recently that several analogs of myristate in which a methylene group is replaced by a single oxygen or sulfur atom are substrates for Saccharomyces cerevisiae and mammalian myristoyl-CoA:protein N-myristoyltransferase (EC 2.3.1.97; NMT) despite their reduced hydrophobicity. Some inhibit HIV-1 replication in acutely infected CD4+H9 cells without accompanying cellular toxicity. To examine the mechanism of their antiviral effects, we performed labeling studies with two analogs, 12-methoxydodecanoate (13-oxamyristate; 13-OxaMyr) and 5-octyloxypentanoate (6-oxamyristate; 6-OxaMyr), the former being much more effective than the latter in blocking virus production. [3H]Myristate and [3H]13-OxaMyr were incorporated into Pr55gag with comparable efficiency when it was coexpressed with S. cerevisiae NMT in Escherichia coli. [3H]6-OxaMyr was not incorporated, even though its substrate properties in vitro were similar to those of 13-OxaMyr and myristate. [3H]13-OxaMyr, but not [3H]6-OxaMyr, was also efficiently incorporated into HIV-1 Pr55gag and nef (negative factor) in chronically infected H9 cells. Analog incorporation produced a redistribution of Pr55gag from membrane to cytosolic fractions and markedly decreased its proteolytic processing by viral protease. 13-OxaMyr and 3'-azido-3'-deoxythymidine (AZT) act synergistically to reduce virus production in acutely infected H9 cells. Unlike AZT, the analog is able to inhibit virus production (up to 70%) in chronically infected H9 cells. Moreover, the inhibitory effect lasts 6-8 days. These results suggest that (i) its mechanism of action is distinct from that of AZT and involves a late step in virus assembly; (ii) the analog may allow reduction in the dose of AZT required to affect viral replication; and (iii) combinations of analog and HIV-1 protease inhibitors may have synergistic effects on the processing of Pr55gag.
Bryant
M L
ML
Department of Pediatrics, Washington University School of Medicine, St. Louis, MO 63110.
Ratner
L
L
Duronio
R J
RJ
Kishore
N S
NS
Devadas
B
B
Adams
S P
SP
Gordon
J I
JI
D000998:Antiviral Agents / Q000494:pharmacology*; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects; D002472:Cell Transformation, Viral; D004926:Escherichia coli / Q000235:genetics; D015683:Gene Products, gag / Q000096:biosynthesis / Q000235:genetics*; D005822:Genetic Vectors; D015497:HIV-1 / Q000187:drug effects / Q000235:genetics / Q000502:physiology*; D006801:Humans; D007700:Kinetics; D007848:Laurates / Q000378:metabolism / Q000494:pharmacology*; D010957:Plasmids; D011498:Protein Precursors / Q000235:genetics; D011499:Protein Processing, Post-Translational; D012441:Saccharomyces cerevisiae / Q000235:genetics; D014779:Virus Replication / Q000187:drug effects; D015215:Zidovudine / Q000494:pharmacology
D000998:Antiviral Agents; D015683:Gene Products, gag; D007848:Laurates; D011498:Protein Precursors; D015215:Zidovudine; C067939:12-methoxydodecanoate