Dear Bruker Users,
I have another question about a 30k Dalton protein on the same AV600 with
cryoprobe.
First I must say that we have run many proteins spectra such as about 8k, 10k ,
20k, 27k on this machine using same pulse programs from biotool. All N-edited
spectra are ok.
I also run one protein with 250 residues, nothing weird happened, However I
run the pro-form of this protein which has additional 25 residues, all things
have been changed. Let me begin from hncacbgpwg3d.
Fist, I can not phase properly using both Bruker standard pulse programs and
nmrpipe. The C dimension can not get pure positive signal of CA and negative
signal of CB.
Second, in the region of H:8.6-7.7, N:125-115, The spectrum is crowded with
very strong, badly phased peaks. Beside this region, we can get some weaker
normal signals, but many signals are missing.The hncacb spectrum is here
http://147.8.148.20/public/yyh/HNCACB.JPG
However, a few months later, the protein degraded to two parts and every part
is about 130 residues. I have run a sds gel to confirm this phenomenon. I run
3D nmr spectra again using the same pulse program and parameters except p1,
p1. I used gpro to input the p1, pl1. All N-edited spectra behave well.
I have checked the 1D proton spectrum without water suppression processing,
the water suppression is very bad.
N-edited spectra, such as hncacb, hncocacb, cbcaconh, hnco, hncaˇK.., all have
these problems for this protein. I have run many times and changed the o1,
swˇK. Every N-edited spectrum, in particular region, that is H:8.6-7.7,
N:125-115 is crowded with badly phased, even hundreds times stronger signals.
Best Regards,
Flora
PhD student
The department of chemistry,
The University of Hong Kong.
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