seleno-methionine

Chris N Larsen clarsen at bimcore.emory.edu
Tue Jan 31 15:23:15 EST 1995


Jon (and others):

Yes, you do put Selenomethionine into proteins the same
way as you do for 35S met. You can grow the bugs up in a defined media, 
with Selenomet instead of met, and as long as the bugs are met-, 
they will make selenoproteins on induction.

I recommend two articles:

MAD phasing of the fibronectin gene:  Leahy et al, 1992. Science 258:987

and (for better methods than Science can provide)

Seleno-xtals of the Gln binding protein:  Hsiao, Cheng-Deng et al. 1994
    J Mol Bio ??:87-92. 

 
The titles are approximate, and sorry, I dont have the issue number,
the net node for this is down. JMB doesnt print them in its crystallography 
notes section.

AND you can get the E coli from STudier and Moffatt or Novagen: 1-800-526-7319. 
Strain B834. They have the DE3 lysogen of this strain too. But if you ask
them how 
to use it, or consult the original cloning paper (ca. 1966) there are no
details. 
So in a pinch, you might go worship the guru of MAD phasing, Wayne 
Hendrickson.

I'VE found that if you grow up the bugs in M9 media (Maniatis) to which 1 mg/ml
Thiamine (yeah, thats right) and Webers "O" and "V" solutions have been added, 
you'll get good growth. Actually, I'm doing the induction right with
Calbiochem's 
SeMET (80$/ 250 mg) and I don't know if it's going to work. You can 
e mail me if you like. The O and V solutions are from David Weber and Albert 
Mildvan, Johns Hopkins University, where they made a 15N protein. Same 
disclaimer applies, but I remember its either Biochemistry or J. Biol. Chem.


GOOD LUCK!!

Chris Larsen




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