Electrostatic repulsion between HIV-1 capsid proteins modulates hexamer plasticity and in vitro assembly.
Capsid protein (CA) is the major component of the human immunodeficiency virus type 1 (HIV-1) core. Three major phosphorylation sites have been identified at positions S(109), S(149) and S(178) in the amino-acid sequence of CA. Here, we investigated the possible consequences of phosphorylation at these sites on the CA hexamer organization and plasticity using in silico approaches. The biological relevance of molecular modeling was then evaluated by analyzing the in vitro assembly properties of bacterially expressed CA bearing S(109)D, S(149)D, or S(178)D substitutions that mimic constitutive phosphorylation at these sites. We found that a constitutive negative charge at position 109 or 149 impaired the capacity of mature CA to assemble in vitro. In vivo, HIV-1 mutants bearing the corresponding mutation showed dramatic alterations of core morphology. At the level of CA hexamer, S(149) phosphorylation generates inter-monomer repulsions, while phosphorylation at position 109 resulted in cleavage of important bonds required for preserving the stability of the edifice. Addition of a negative charge at position 178 allowed efficient assembly of CA into core-like structures in vitro and in vivo and significantly increased CA hexamer stability when modeled in silico. All mutant viruses studied lacked infectivity since they were unable to produce proviral DNA. Altogether our data indicate that negative charges, that mimic phosphorylation, modulate assembling capacity of CA and affect structural properties of CA hexamers and of HIV-1 cores. In the context of the assembled core, phosphorylation at these sites may be considered as an event interfering with core organization and HIV-1 replicative cycle.
Brun
Sonia
S
Centre d'études d'agents Pathogènes et Biotechnologies pour la Santé (CPBS), Université Montpellier 1, Montpellier, France.
Chaloin
Laurent
L
Gay
Bernard
B
Bernard
Eric
E
Devaux
Christian
C
Lionne
Corinne
C
Chazal
Nathalie
N
Briant
Laurence
L
D036022:Capsid Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D045744:Cell Line, Tumor; D015497:HIV-1 / Q000235:genetics*; D006801:Humans; D056004:Molecular Dynamics Simulation; D009154:Mutation; D010641:Phenotype; D010766:Phosphorylation; D055503:Protein Multimerization; D055550:Protein Stability; D055672:Static Electricity; D014771:Virion; D054301:gag Gene Products, Human Immunodeficiency Virus / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism
D036022:Capsid Proteins; D054301:gag Gene Products, Human Immunodeficiency Virus