Hi,
Generally speaking, these days you don't need a special vector to introduce a C-His (or an N-His, BAP, STREP, STREPII, or any short tag for that matter), since the sequence of the tag + cleavage site is short enough to be engineered directly into your PCR amplicon. There are numerous ways to cleave C-terminal tags, however most of them leave several residues behind, due to the specificity determinants of (most) proteases being upstream of the cut site. If you absolutely have to use C-terminal His, consider not removing it at all since the difference between a site-specific protease 'leftover' and a 6-His stretch isn't very significant (although of course the effect on crystallization can be profound!). There are several somewhat esoteric ways to get rid of His-tags at the C-terminus cleanly - these involve treatment with carboxypeptidases and other similar tricks. Artem _____ From: CCP4 bulletin board [mailto:[EMAIL PROTECTED] On Behalf Of Vijay Kumar Sent: Friday, April 04, 2008 11:40 AM To: [email protected] Subject: [ccp4bb] off topic - C-ter cleavable his tag Hi, I am looking for a vector with a cleavable C-terminal his tag for prokaryotic protein expression. I found pET-52b but it has also got a N-terminal cleavable strep-tag which I cannot avoid. Thanks Vijay
