Universal 13C-Asp labelling of Protein

John Tomaszewski tomasz at protein.chem.washington.edu
Tue May 13 20:05:13 EST 1997

	We have overexpressed a small (15KDa) protein and a couple of 
mutants in both LB and minimal media (for 15N labelling) in our lab but 
now we have a need to express it (and one mutant) universally 13C-Asp 
labelled.  What I'm trying to hear are peoples experiences with some of 
the options along these lines.
	It seems to me that it might be possible with either LB or 
minimal media seeing as how ~25% 13C enrichment should be plenty for 
13C NMR.  Some of my questions are:
1) Can we simply spike LB with 13C-Asp and achieve the desired level 
of enrichment or will the 13C-Asp be broken down and the 13C scattered 
around too fast?
2) Would minimal media with all AAs (and only 13C-Asp) supplied be a 
better way to go?
3) What is the metabolic fate of the 13C in 13C-Asp?
4) Would be worth our effort to transfer the plasmid to the appropriate 
auxotroph and go from there?

	Any help, suggestions, pointers to papers, etc. would be greatly 
appreciated.  Thank you.


John T.

**  **  ** ** * **            DEPARTMENT OF CHEMISTRY             **
**  **  ** ** * **        tomasz at protein.chem.washington.edu      **
**  **  ** ** * **                (206) 616-2993                  **
**   *****  *****                JOHN TOMASZEWSKI                 ** 

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