Research methods in protein-protein and protein-nucleic acid interactions and application in the study of human enterovirus A71 / 病毒学报

Methods for analyses of protein-protein interactions include: yeast two hybrid (Y2H), phage dis- play (PD), co-immunoprecipitation (Co-IP), glutathione S-transferase pull-down (GST pull-down), cellular co-localization, far-western blotting, virus overlay protein binding assay (VOPBA), surface plasmon resonance (SPR), and fluorescence resonance energy transfer (FRET). Technologies for the detection of protein-nucleic acid interactions include: yeast one hybrid (Y1H), chromatin immunoprecipitation (ChIP), electrophoretic mobility shift assay (EMSA), Southwestern blotting, reporter gene, Co-IP, GST pull-down, and PD. These methods are often used in the study of the human enterovirus A71 (EV-A71) by our research team. Reviews in the Chinese literature in this field are lacking, so we reviewed applications of these methods in the study of EV-A71. This review may impart important knowledge in the research of other viruses with regard to protein-protein and protein-nucleic acid interactions.

Epitope analysis of cDNA sequences of Deinagkistrodon acutus snake venom metalloproteinases and observation of their immune protective effects / 第二军医大学学报

Objective: To analyze the epitopes of cDNA sequences of Deinagkistrodon acutus snake venom metalloproteinases using bioinformatical method, and to observe the immune protective effect of the new immunogen designed according to the identified epitopes. Methods: The cDNA sequences of Deinagkistrodon acutus snake venom metalloproteinases were amplified by PCR. The epitopes of the sequences were analyzed by Jameson-Wolf method and Clustal X software, then the sequences of the screened epitopes were artificially synthesized and linked to the vector pIRESneo. BALB/c mice were immunized by the resultant plasmid at 0, 2, and 4 weeks for three times, then the titers of the anti-serum were measured by ELISA. The immune protective effects of the anti-serum were tested by the neutralization of venom hemorrhagic activity and venom attacking test. Results: Bioinformatical analysis yielded 6 epitopes ( MPA-1-MPA-6). The ELISA results of anti-serum showed that these epitopes could induce immune reaction in mice, and the anti-serum was detectable even when it was diluted to 1:100. The neutralization test and venom attacking test demonstrated that the anti-serum induced by the epitodes could neutralize the venom and protect the mice from haemorrhage. Conclusion: Six epitopes of Deinagkistrodon acutus snake venom metalloproteinases have been obtained successfully using bioinformatical method, and the new immunogen designed based on these epitopes shows a primary immune protective effect.