Affinity modification of tryptophanyl-tRNA synthetase by an alkylating L-tryptophan analog].

G K Kovaleva, S Kh Degtiarev, O O Favorova

Molekuliarnaia biologiia 1979 Nov-Dec

filter terms:

3-Amino-1-chloro-indolwbutan-2-one (Trp-CH2Cl) was synthesized to be used for labeling the active site of tryptophanyl-tRNA-synthetase. Trp-CH2Cl irreversibly inhibits the beef pancreas tryptophanyl-tRNA synthetase activity. The inhibition rate was found to exhibit saturation concentration dependence typical for an affinity reagent. L-tryptophan and L-tryptophanyl adenylate protect the enzyme from inhibition. To determine the stoichiometry of inhibitor--protein binding 3H-label from NaB3H4 was incorporated into the modified enzyme. The molar ratio of inhibitor residues incorporated into the modified enzyme (dimeric molecule) is approximately 2. When one of the subunits of the enzyme was reversibly protected with relatively stable tryptophanyl adenylate, the modification of this enzyme led to the blocking of the other subunit (so called "one-site" enzyme). Some properties of the "one-site" enzyme obtained were studied.

Citation

G K Kovaleva, S Kh Degtiarev, O O Favorova. Affinity modification of tryptophanyl-tRNA synthetase by an alkylating L-tryptophan analog]. Molekuliarnaia biologiia. 1979 Nov-Dec;13(6):1237-46