Partial cleavage of HA (Hemagglutinin) tag? - (Feb/20/2013 )
I'm expressing a protein with an N-terminal HA tag which I am using for identification purposes.
A quality-control LC-MS suggests that my protein has 2 populations composed of: 1) "full-length HA tag + protein" and 2) "partially cleaved HA tag + protein"
No other cleavage appears to be evident within my protein since the observed MW agrees closely with the theoretical MW.
Ps. Protein is expressed in CHO cells and is secreted into the media.
Expression construct consists of: 'Signal peptide + HA tag + protein'
Are you sure that it is cleavage and not skipped expression?
I was thinking of a second ATG other than that I don't think skipped expression is possible.
Which enzyme do you use before MS? If you use Chymotrypsin, your HA tag is highly possible to be cleaved. Check your HA sequence.