Perparing a Selenomethionine derivative
for a MAD experiment

From Robert Frank Standaert's Thesis: Consultation of a biochemistry textbook indicated that threonine, isoleucine, and lysine inhibit methionine biosynthesis in E. coli by inhibiting aspartokinases, and that phenylalanine and leucine are synergistic with lysine.  Incorporation of this information into a strategy for SeMet labeling was simple. E. coli XA90 were grown in 1 L of M9 medium to mid-log phase (A600=0.6), and 60 mg of SeMet (sigma) along with 100 mg each of threonine, lysine hydrochloride, and phenylalanine and 50 mg each of leucine, isoleucine, and valine were added as solids to the growing culture.  After another 15 min, IPTG was added to 1 mM, and the procedure was completed as usual.  Purification was performed essentially as for native protein, except that DTT at a concentration of 5-10 mM was maintained throughout on the expectation that SeMet would be sensitive to oxidation. The yield of protein was 10 mg, about what would be obtained from a normal prep. 

I used to buy my SeMet from Acros in 1998.
 It was 10 times cheaper than Sigma and it works.


J. Mol. Biol (1993) 229, 105-124, 1993


[Overview] ·[Facilities] · [People] · [Services] ·[Lectures] · [BioLinks] · [Stats] ·[Search]