Site-specific isotopically-labeled proteins, amino acids, and biochemical precursors therefor

Abstract

Site-specific isotopically-labeled valine, leucine, and isoleucine and biosynthetic precursors for these amino acids are provided. The amino acids are labeled with 13 C or 14 C at the methyl group carbon atom(s) most remote from the carboxyl group. Also disclosed are the biochemical precursors of these labeled amino acids, 2-keto-4-( n C) butyric acid and 2-keto-3-( n C-methyl)-4-( n C)-butyric acid in which n, at each occurrence, is 13 or 14. Also disclosed are proteins, protein fragments, and polypeptides containing these site-specifically isotopically labeled amino acids, and methods for preparing the biochemical precursors, the amino acids, and the proteins, protein fragments, and polypeptides.

1 - 11 . (canceled)  
     
     
         12 . A method of preparing a protein, protein fragment, or polypeptide containing at least one amino acyl residue selected from the group consisting of 4-( n C-methyl)-5-( n C)-leucyl, 5-( n C) n C)-isoleucyl, and 3-( n C-methyl)-4-( n C)-valyl, which comprises 
 a) genetically modifying a suitable microorganism to express said protein, protein fragment, or polypeptide;    b) culturing said genetically modified microorganism in a nutrient medium containing a compound of formula Ia                          or a salt thereof, wherein n, at each occurrence is 13 or 14, and R 3  is hydrogen or  n CH 3 ; and    c) isolating said protein, protein fragment, or polypeptide.    
     
     
         13 - 16 . (canceled)