Functional Surfaces of the Human Immunodeficiency Virus Type 1 Capsid Protein

Abstract

The human immunodeficiency virus type 1 initially assembles and buds as an immature particle that is organized by the viral Gag polyprotein. Gag is then proteolyzed to produce the smaller capsid protein CA, which forms the central conical capsid that surrounds the RNA genome in the mature, infectious virus. To define CA surfaces that function at different stages of the viral life cycle, a total of 48 different alanine-scanning surface mutations in CA were tested for their effects on Gag protein expression, processing, particle production and morphology, capsid assembly, and infectivity. The 27 detrimental mutations fall into three classes: 13 mutations significantly diminished or altered particle production, 9 mutations failed to assemble normal capsids, and 5 mutations supported normal viral assembly but were nevertheless reduced more than 20-fold in infectivity. The locations of the assembly-defective mutations implicate three different CA surfaces in immature particle assembly: one surface encompasses helices 4 to 6 in the CA N-terminal domain (NTD), a second surrounds the crystallographically defined CA dimer interface in the C-terminal domain (CTD), and a third surrounds the loop preceding helix 8 at the base of the CTD. Mature capsid formation required a distinct surface encompassing helices 1 to 3 in the NTD, in good agreement with a recent structural model for the viral capsid. Finally, the identification of replication-defective mutants with normal viral assembly phenotypes indicates that CA also performs important nonstructural functions at early stages of the viral life cycle.

In Vitro Identification and Characterization of an Early Complex Linking HIV-1 Genomic RNA Recognition and Pr55Gag Multimerization

“…The process of multimerization and assembly can be seen as a protein folding process in which two alternatives are possible: proper folding and assembly versus aggregation. The M318A mutation strongly reduced HIV-1 infectivity with reduced particle production and defective assembly of capsid and Gag (71). Our data suggest that the early association of two Gag proteins with the 5Ј-LR provides a conformation that is crucial for assembly and proper Gag-Gag interactions.…”

“…SL3 & SL2) and the C-terminal domain of CA has a dimer interface. The presence of a M318A mutation, which is known to impair each of CTD dimerization, proper Gag assembly, and viral infectivity (23,71), also abolished formation of the EC. The EC apparently has an additive impact on the affinity of protein for genomic RNA since M318A has approximately half of the binding affinity of wt mGag ([K d ] ϭ K d ⅐n) but was crucial for multimerization ability.…”

Structural and Dynamics Studies of the D54A Mutant of Human T Cell Leukemia Virus-1 Capsid Protein

“…Residues with NOE Ͻ 0.65 and residues that exhibit conformational exchange on a microsecond/millisecond time scale must be excluded. We found 17 residues with NOE values lower than 0.65 (Met 3 133 , and Ala 134 ). The same criteria used to identify the WT residues with slower internal motions (16) were used in evaluation of the mutant.…”

“…The mature virion is conical for human immunodeficiency virus type 1 (HIV-1) and other lentiviruses and spherical or irregularly polyhedral for human T cell leukemia virus type 1 (HTLV-1) and other members of the retrovirus group, including Rous sarcoma virus and Moloney murine leukemia virus. The morphology of the core and viral infectivity are closely correlated; mutations that result in loss of the characteristic core structure also result in loss of infectivity (2,3).…”

“…The structure is highly conserved among retroviral CA proteins that have been examined (5)(6)(7)(8)(9)(10); mutations in the C-terminal domain of HIV-1 block CA protein dimerization and particle assembly (3,(11)(12)(13). Mutations in the N-terminal domain (NTD) of HIV-1 impair CA protein assembly to a significantly lesser extent, but the assembled particles are noninfectious (3,11,13,14). In contrast, mutations in the C-terminal domain of the HTLV-1 CA protein had little effect on viral particle assembly and release, whereas mutations throughout the NTD of the protein had much greater impact (15).…”