Résumé
APOBEC3 C is special as it has only weak antiviral functions and weakly restricts retroelements compared to other APOBEC3 s. APOBEC3 has only one cytidine deaminase domain which coordinates a zinc ion, and then is classified to A3 Z2 according to the amino acid specificity. APOBEC3 C induces less cytidine deamination in HIV-1 DNA than APOBEC3 G and has reduced ability to inhibit HIV-1 reverse transcription and integration compared to APOBEC3 G. APOBEC3 C induces G-to-A mutation that cannot block viral replications but contribute more to viral diversity. A single nucleotide polymorphism in human APOBEC3 C, a change from serine to isoleucine at position 188, results in increased enzymatic activity and potent antiviral activity against HIV-1. Dimerization of human APOBEC3 C increases the ability of continuous synthesis of single-stranded DNA, resulting in higher levels of mutation during reverse transcription in vitro and in cells. APOBEC3 C has evolved under positive selection in primates, and it is an important barrier that must be countered by the virus during natural infections. Therefore, research on APOBEC3 C may provide some ideas for anti-retroviral and anti-cancer therapeutic design.
Résumé
Apolipoprotein B mRNA editing enzyme-catalytic polypeptidelike 3G (APOBEC3G), which belongs to the APOBEC family, restricts the replication of HIV-1 through the deminase-dependent and deminase-independent antiviral mechanisms. Viral infectivity factor (Vif), an accessory protein edited by vif gene of HIV-1, promotes APOBEC3G ubiquitination and subsequent degradation by the proteasome result ing in the increased replication of HIV-1. Over the past few years, the discovery of some co-factors such as CBF-β, HDAC6 and MDM2 not only raises our awareness of the interaction between APOBEC3G and Vif but also provides new antiviral targets of HIV-1 in addition. Here we review the structure of APOBEC3G and Vif, and the effect of interference agents on HIV-1 replication by interacting with APOBEC3G and Vif.
Résumé
OBJECTIVE: To design and synthesize HIV-1 Vif inhibitor RN-18 derivatives and investigate their antiviral activities. METHODS: RN-18 was used as the lead compound, and optimizations were carried out on the two side chains and the middle benzene ring. The anti-HIV-1 activities of the target compounds were analyzed by p24 assay, and the cytotoxicities of compounds with better activities were tested with MTT method. RESULTS: Twenty-eight new derivatives were prepared, and their structures were characterized by MS and 1H-NMR. The activity test showed that the antiviral activities of seven compounds were obviously improved, and the activity of compound 26 was increased to be 11 times higher than that of RN-18, with an EC50 value of 21.8 μmol·L-1. CONCLUSION: Replacing the middle benzene ring of RN-18 with heterocycles generally improves the anti-HIV-1 activity, which provides the basis for further study.
Résumé
Human APOBEC3G (hA3G) is a cytidine deaminase which inhibits HIV-1 replication. The HIV-1 accessory protein viral infectivity factor (Vif) counteracts with hA3G by targeting it for proteasomal degradation. In this work, we constructed and optimized molecular models of the hA3G dimer and the hA3G-Vif complex. The molecular modeling study revealed that the loop7 motif of hA3G appears on the interfaces of both the hA3G-Vif complex and the hA3G dimer. Biochemical analysis provided evidence suggesting that binding of Vif to hA3G results in steric blocking of hA3G dimerization, implying that monomeric hA3G serves as a substrate for Vif-mediated degradation. Furthermore, we presented evidence for the important roles of the loop7 motif, especially the central residues within the region, in hA3G dimerization, hA3G--Vif interaction, Vif-mediated hA3G degradation as well as subcellular localization of hA3G. This work highlights a multiple-task interface formed by loop7 motif, which regulates biological function of hA3G, thus providing the feasibility of the strategy of blocking Vif-mediated A3G degradation by targeting the putative site around loop7.
Résumé
Interaction between the HIV-1 Vif protein and the cellular host APOBEC3G protein is a promising target for inhibition of HIV-1 replication. Considering that human cells are a very complicated environment for the study of protein interactions, the goal of this study was to check whether fission yeast could be used as a model cell for studying the Vif-APOBEC3G interaction. Vif and APOBEC3G were expressed in fusion with GFP protein in the S. pombe SP223 strain. Subcellular localizations of Vif and APOBEC3G were observed with fluorescent microscopy. Codon optimization was used to over express the Vif protein in S. pombe cells. The degradation of APOBEC3G mediated by Vif was tested through expressing Vif and GFP-APOBEC3G proteins in the same cell. Western Blot analysis was used to measure the corresponding protein levels under different experimental conditions. The results showed that the Vif protein was predominantly localized in the nucleus of S.pombe cells, APOBEC3G was localized in the cytoplasm and concentrated at punctate bodies that were often in close proximity to the nucleus but were not necessarily restricted from other regions in the cytoplasm. Vif protein expression levels were increased significantly by using codon optimization and APOBEC3G was degraded when Vif was over-expressed in the same S. pombe cells. These results indicate that fission yeast is a good model for studying the interaction between the Vif and APOBEC3G proteins.
Résumé
Objectives To analyze the characteristic of HIV-1 viral infectivity factor (Vif) gene variants isolated from Shanghai. To construct the prokaryotic expression vector of HIV-1 vif gene and understand its immunogenieity. Methods HIV-1 vii genes were amplified and sequenced from 23 serum samples of HIV-1 infected patients in Shanghai and then compared with the international standard HIV-1 strain. Subsequently, these amplified Vif fragments were sub cloned into pETS2b(+)expression vector. The recombinant prokaryotie plasmids pETS2b (+)-HIV-1/Vif were then transferred into BL21DS(Star)cells for expressing and purifying HIV-1 Vif protein. HIV-1 Viff rat polyclonal antibody was then preparing by injecting the purified Vif proteins into the mice. ELISA was used to determine the purity of Vif proteins and the immunogenicity of its polyclonal antibodies. Results The nucleotide acid mutation rate of HIV-1 vif gene in Shanghai AIDS patient was (0. 179±0. 006)% compared with the international standard HIV-1 strain. Some similar mutations in vif gene were found in HIV-1 strains isolated from Shanghai while the amino acid sequence between 151 and 240 in Vif protein was conserved. The construction of HIV-1 Vif prokaryotic expression plamids and the preparation of Vif polyclonal antibodies were successfully done in this study. The reaction between recombinant HIV-1 Vif protein and the serum from HIV-1 infected patient was not significantly different from that between the recombinant protein and healthy control serum(P＞0.05). HIV-1 Vif polyclonal antibody reacted differently with recombined HIV-1 Vif protein compared with healthy control serum samples (t=178.61, P＜0.01). Conclusions The Vif gene mutation rate is high in HIV-1 strains isolated from Shanghai compared with international standard HIV-1 strain. The prokaryotic expression plasmids of HIV-1 vif antigen are successfully constructed and Vif polyclonal antibodies are prepared well.