Hey, I am characterizing a competitive inhibition of a host and viral RNA biding proteins. The RNA binding is not sequence specific but dependent on secondary structure of the RNA. I am looking for a method to identify if the viral protein blocks the host protein from binding RNA in cells.
Any suggestions would be really helpful but I was also specifically considering a filter binding assay or EMSA but I can not isotope label the RNA in my lab. Though I would check if anyone had suggestions.
Thanks for your help!
Hi Heather,
My idea may be too naive: I guess your proteins are generated in cells or in vitro. Can't you just produce them with individual tags, then do a pull-down and quantify the RNA?
If you are able to label your RNA of interest then competitive RNA -EMSA is method of choice to see preferential binding between host protein or Viral RBP.In addition you can also perform RIP (RNA immunoprecipitation) , Biotin RNA pulldown etc to validate competition or inhibition.
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