Dear Crystallographers (this may be off-topic, depending on whom you ask...)
A: Can anyone recommend new improved alternatives to the usual suspects for proteases (TEV, thrombin, enterokinase, etc.)? I've seen some literature about SUMO- and NEDD8-dependent enzymes, but those apparently require the whole protein domain to be in the construct for cleavage to happen, rather than just a short sequence motif. Further, I'd be curious whether there be drawbacks to using the various new breeds that might not be mentioned in the original publications thereon. B: While I have seen several proteases that leave behind only 1 aa on the new n-terminus of the target, I've yet to come across proteases that leave behind very few residues on the new c-terminus, which would be very helpful for tagging cleanly the c-terminus of proteins. Do such enzymes exist, and if so, are there particularly good ones? And if not, I wonder why proteases usually require more sequence on the n-terminal side of the scissile bond? All the best, Jacob Keller ******************************************* Jacob Pearson Keller, PhD Looger Lab/HHMI Janelia Farms Research Campus 19700 Helix Dr, Ashburn, VA 20147 email: kell...@janelia.hhmi.org *******************************************
