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When pursuing an NMR structure project - at what size of oligopeptide you would decide to label it with:
Have you worked with a non-labeled oligopeptide where you thought later "I wish I've labeled it" - how many residues did it have? Thanks. |
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Your title says protein, but your post says oligopeptide. If you're synthesizing peptides, cost can be a real issue. I recently did a pair of 7-mers unlabeled from NOESY, COSY, and TOCSY and it wasn't bad once I got into it, but all the spin systems were relatively unique. I've also assigned most of an unlabeled 24-mer in a protein-peptide complex with multiple non-unique amino acids using filtered 2D NOESY and TOCSY, and that was MUCH more difficult, but still possible. With actual protein, assuming you can express in bacteria or yeast, my suggestion would be to make an initial batch 15N only to optimize sample conditions and make sure the purified protein is well behaved - you can use this same sample for any 15N-NOESY or relaxation data you want to collect. Once you've gotten conditions worked out, there's absolutely no reason not to use a double labeled sample these days - the extra cost of isotope and a few more 3Ds on the instrument will more than pay for themselves in time savings and fewer errors in assignment. Thanks Andrew! I've corrected the title a bit, well I guess there is a fine line somewhere in terminology :). 24-mer peptide in the complex is quite a feat! - Evgeny Fadeev (Jul 30 '10 at 08:08) I would always suggest to label if possible. So much more you can do. One way is to express the peptide as C-terminal tag in, say, a GB1-TEVsite-fusion protein that can be subsequently cleaved by TEV protease. There are other ways to do this where the peptide is directed to inclusion bodies. - Paul Driscoll (Feb 02 '11 at 04:28) |
