I have infected insect cells SF9 with recomb. baculovirus expressing Core.His and another with GFP.
I have extracted the soluble proteins after 3 days and did western blot using anti-his, anti-GFP and anti-tubulin.
the GFP and tubulin gave clear bands at right size but nothing for the Core-His.
I am wondering if I should have extracted insoluble proteins from the pellet? and if so what is the recommended protocol?
Years ago, I worked on the HBV core protein. If the HCV one is any similar to its HBV cousin, you will have a tough time ahead.
I am pretty sure that you will find it in the insoluble fraction. We needed a mixture of both urea and guanidine HCl to get the HBV core protein extracted.
Your core protein is probably highly hydrophobic and practically insoluble in aqueous solutions. I assume you have the protein sequence; check it for its hydrophobicity. We kind of got the HBV core soluble, but not as individual protein, but as a micellar structure and only at very acidic pH-values that made it difficult to use for in vivo studies.
Thanks Achim. I checked the sequence and I found it mostly consists of hydrophobic amino acids, will try to extract with that mixture and hopefully it works.
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