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Mutagenesis studies of the PTAPP sequence in the HIV-1 p6Gag protein.


Natl Conf Hum Retroviruses Relat Infect (1st). 1993 Dec 12-16;:127.

Point mutations were introduced into a full length HIV-1 proviral clone (pNL4-3) to investigate the biological significance of a conserved amino acid sequence (PTAPP). This sequence is located near the C-terminus of Gag precursors in most lentiviruses and all HIV-1 strains. Mutants containing the following amino acid substitutions for the PTAPP coding sequence were constructed: LTAPP, PTALP, PTAPL, PAAPP, PTNPP, and LTALL. Effects on full- length genomic RNA content and viral infectivity were determined. Upon transfection into HeLa cells, all the mutant proviruses produced virus-like particles with properly processed p7NC, p24CA, gp120SU and functional RT and Pr. The LTALL mutant produced approximately 20-fold less virus than wild-type and the other mutants. Mutant viral stocks, normalized on the basis of p24CA content, were analyzed for infectivity in H9 cells and for genomic RNA by Northern analysis. The infectivities ranged from near wild- type levels to non-infectious in the following order: wild- type greater than or equal to PTAPL, PAAPP greater than LTAPP, PTALP, PTNPP greater than LTALL. In most cases, the reduction of mutant virus infectious titer appeared to parallel the reduction in full length genomic RNA content. The data suggest that single amino acid substitutions in the PTAPP sequence of the Gag Precursor (p6 domain) can reduce the efficiency of genomic RNA packaging or protection without influencing particle production or infectivity. However, multiple mutations in this region (LTALL) can result in the production of non-infectious, RNA-deficient particles, similar to those observed from HIV-1 ZN++-finger mutants.

Amino Acid Sequence Blotting, Northern Gene Products, gag/*GENETICS Hela Cells Human HIV Protease/GENETICS HIV-1/ENZYMOLOGY/*GENETICS Molecular Sequence Data *Mutagenesis RNA-Directed DNA Polymerase/GENETICS Transfection ABSTRACT


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