i like this post (click again to cancel)
1
i dont like this post (click again to cancel) remove favorite mark from this question (click again to restore mark)

Hi I am observing some unexpected cross peaks in some homonuclear protein data and I was wanting to know if anyone had an ideas what im looking at. I am looking at what I was confident was a tyrosine HE-HD cross peak in a TOCSY giving charachterticaly intense cross peaks at 6.9 x 6.4 ppm, however I then noticed a second set of TOCSY peaks to both the HE* and HD* at 5.4 ppm which also gives a COSY to the HE. These peaks shift together if I change the temperature sugesting it is not just two overlaped resonances, are still present in D2O, both the HE and HD* give the anticipated intensity in the 1D sugesting theses are not chemical exchange cross peaks. my initial gues was observation og the Tyr OH but the chemical shift doesnt really make sense and one wouldnt expect to see a COSY to the OH either, this is not ovrlaped with an NH2 or NH (by compsrison with my HSQC) any ideas, im at a bit of a loss and not sure if im just not missing an obvious explanation. Any help would be apreciated Tom

asked Oct 16 '11 at 20:59

Thomas%20Garner's gravatar image

Thomas Garner
81


3 Answers:
i like this answer (click again to cancel)
0
i dont like this answer (click again to cancel)

Your description of the data, although not entirely clear, appears to suggest a phenyl-alanine rather than a tyrosine. You should check your assignments first.

A triple quantum filter will remove the cross-peak if it involves HD-HE of tyrosine, but will not do so if it is from phenyl-alanine. Therefore, a triple quantum filtered 2D COSY should distinguish between these two possibilities.

You cannot rule out the possibility that the amino acid sequence of the protein has been determined incorrectly - I know of a case where NMR data was used in the course of protein structure determination by Prof. Wagner to correct a mistake in the amino acid sequence of the protein.

However, the (very unlikely) possibility of a Tyrosine OH cannot be ruled out inspite of the unusual chemical shift and the presence of cross peaks in D2O. If the Tyrosine OH is buried and hydrogen bonded, then mild heating in D2O may not lead to exchange with solvent D2O. If your protein can be completely unfolded by heating in D2O, and then renatured by cooling, then the cross peak should dissappear from D2O COSY spectrum if the cross peak involves a J coupling between Tyrosine OH-HE.

Note: The thermal denaturation and renaturation should be carried out in a dilute solution of the protein in D2O, to prevent aggregation.

link

answered Dec 03 '11 at 12:55

sekhar%20Talluri's gravatar image

sekhar Talluri
621

updated Dec 03 '11 at 12:57

i like this answer (click again to cancel)
0
i dont like this answer (click again to cancel)

L-Tyrosine (CAS No.60-18-4, HS code292250) is a amino acid manufactured through fermentation, available as White crystals or crystalline tasteless powder. L-Tyrosine is widely used as nutrition supplements. It is widely accepted as safe food additive in many countries.

link

answered Aug 01 '16 at 21:28

james%20han's gravatar image

james han
1

i like this answer (click again to cancel)
0
i dont like this answer (click again to cancel)

I know a commercial platform that provides custom protein services, such as crystallization, structure determination and function analysis by X-ray, EM, NMR, etc.

link

answered Jun 27 '16 at 22:42

Justin%20Frank's gravatar image

Justin Frank
1

Your answer
Please start posting your answer anonymously - your answer will be saved within the current session and published after you log in or create a new account. Please try to give a good answer, for discussions, please use comments and please do remember to vote (login to vote)
toggle preview

Tags:

×2
×1

Asked: Oct 16 '11 at 20:59

Seen: 2,733 times

Last updated: Aug 01 '16 at 21:28

powered by CNPROG