Study of humoral immune responses of a mutant within CD4 binding site of human immunodeficiency virus type 1, envelop glycoprotein in mice.

Abstract

Objective To investigate the effect of the triple amino acid residues I423, N425 and G431 within CD4 binding site core region (CD4BScore) of human immunodeficiency virus type 1 (HIV-1) envelope glycoprotein (Env) on the Env immunogenicity, the humoral immune responses of an envelope gp120T-ING/MKE mutant was evaluated in BALB/c mice. Methods Thegp120 gene mutant ING/MKE at I423, N425 and G431 within gp120 CD4 binding site of HIV-1 06044 strain was PCR amplified. The recombinant plasmid pcT22-06044gp120T-ING/MKE (gp120T-ING/MKE) was constructed. HEK293T cells were transiently transfected with the recombinant plasmids to express the wild type (gp120Twt) and the mutant gp120 (gp120T-ING/MKE) trimeric proteins. BALB/c mice were immunized using DNA prime and protein boost regimen. Fourteen days after the last immunization, the binding and neutralizing antibody responses in mouse sera were measured by using an enzyme immunoassay and HIV-1 pseudovirus neutralization assay in TZM-bl cells. Antibody-secreting plasma cells in bone marrows were also detected by using B cell ELISPOT. Results The plasmid gp120T-ING/MKE was constructed and confirmed by DNA sequencing. The recombinant trimeric gp120 proteins were expressed in the culture supernatants. Day14 after the final immunization, sera from the immunized mice by gp120Twt and the mutant exhibited similar specific binding activities. The mutant-immunized mice had lower frequencies of specific and non-specific antibody-secreting cells than gp120Twt-immunized mice. Both gp120Twt and ING/MKE mutant induced weak neutralizing activities against SF162 pseudoviruses. Conclusion The triple mutation ING/MKE in CD4BS did not improve the immunogenicity of gp120. Key words: Human immunodeficiency virus; Envelope glycoprotein; CD4 binding site; Mutation; Immunogenicity