I am purifying a protein with a his tag. The TEV cleavage site I read is composed of ENLYFQG, and this G remains after cleavage. What I found to also remain is an 'am'. It is before the first sequence in the N terminal domain of my protein. Can anyone explain what this is? Is it actually amino acids in my protein sequence or could it be something remaining from the cleavage? I see a clear density for these 3 amino acids 'gam' in my xray crystal structures.
Thanks !
You need to verify your target gene from the expression construct by sequencing. The sequence might have been added during cloning by mutation. Does the "am" sequence have significant effect on the physico-chemical properties or functionalities of your target protein? Although it is good to identify the root causes that added the sequences to the protein, I will not bother about it provided that the physico-chemical properties and functionalities of the protein are not affected because of the presence of "am" at the N-terminal.
Hi there,
What was your cloning strategy (vector, restriction enzymes, primer sequences)? I suspect the AM sequence comes from the vector...
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