Detection of a trimeric human immunodeficiency virus type 1 Gag intermediate is dependent on sequences in the matrix protein, p17.
Previous studies have shown that single amino acid changes in the amino-terminal matrix (MA) domain, p17, of the human immunodeficiency virus type 1 Gag precursor Pr55, can abrogate virion particle assembly. In the three-dimensional structure of MA such mutations lie in a single helix spanning residues 54 to 68, suggesting a key role for this helix in the assembly process. The fundamental nature of this involvement, however, remains poorly understood. In the present study, the essential features of the MA helix required for virus assembly have been investigated through the analysis of a further 15 site-directed mutants. With previous mutants that failed to assemble, residues mapped as critical for assembly were all located on the hydrophobic face of the helix and had a key role in stabilizing the trimeric interface. This implies a role for the MA trimer in virus assembly. We support this interpretation by showing that purified MA is trimeric in solution and that mutations that prevent virus assembly also prevent trimerization. Trimerization in solution was also a property of a larger MA-capsid (CA) Gag molecule, while under the same conditions CA only was a monomer. These data suggest that Gag trimerization driven by the MA domain is an intermediate stage in normal virion assembly and that it relies, in turn, on an MA conformation dependent on the hydrophobic core of the molecule.
The Kitasato Institute, Minato-ku, Tokyo 108, Japan. ymorikawa@kitasato.or.jp
Y
Y
Morikawa
W H
WH
Zhang
D J
DJ
Hockley
M V
MV
Nermut
I M
IM
Jones
D000818:Animals; D002460:Cell Line; D015870:Gene Expression; D015683:Gene Products, gag / Q000235:genetics* / Q000378:metabolism; D015488:HIV Antigens / Q000235:genetics* / Q000378:metabolism; D015497:HIV-1 / Q000235:genetics* / Q000502:physiology*; D006801:Humans; D016296:Mutagenesis; D011498:Protein Precursors / Q000235:genetics* / Q000378:metabolism; D012996:Solutions; D018411:Spodoptera; D014764:Viral Proteins; D014771:Virion; D019065:Virus Assembly; D054301:gag Gene Products, Human Immunodeficiency Virus
D015683:Gene Products, gag; D015488:HIV Antigens; D011498:Protein Precursors; D012996:Solutions; D014764:Viral Proteins; D054301:gag Gene Products, Human Immunodeficiency Virus; C054858:p17 protein, Human Immunodeficiency Virus Type 1; C059767:p55 gag precursor protein, Human immunodeficiency virus 1