The virus family on which I work, has separate fusion protein and receptor attachment protein, I have two subtypes of this virus, one has shorter fusion protein (highly pathogenic) and other has longer fusion protein ( weak pathogen), is there any assay I can understand how these fusion protein length can affect viral replication capacity,
Hi Sushant, site directed mutagenesis to alter the position of the fusion stop codon followed by Transfection and virus recovery ,then application of invitro and invivo pathogenicity experiments. this trial will emphasis the role of fusion protein length in pathogenicity.
regards
Dear Sushant - in case of your virus, is pathogenicity in vivo in good correlation with in vitro replication kinetics? Are there any other molecular/protein differences between subtypes which can interfere with the experiment? Is cellular receptor identified (if so, you can try with receptor-ligand inhibition). Ehab's idea looks great, especially if you are able to identify interdomain sequence (assuming hypothesis that there are two different fusion domains in two virus subtypes). If changes in fusion kinetcs are because of differences in single domain present in both subtypes, I would start with in silica analysis of the domains sequences.
Maybe changes in fusion ability can be identified in other way - if the virus causes syncytial CPE - differences in viral fusion proteins could result in efficacy of syncytia formation, that could be good for start.
you may find information in the links below:
Article Drug Resistance Mutation Profile and Accumulation Kinetics i...
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4705996/
https://pdfs.semanticscholar.org/cd36/9d4a648ce391b8ab563c2a8e6651ced85295.pdf
Plaque assays are the most reliable method of determining infection kinetics.
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