Post-translational modification of a tryptophan biosynthetic enzyme. Precursor-product relationships in vivo and...

Purified preparations of the bifunctional, tryptophan biosynthetic enzyme N-(5'-phosphoribosyl)anthranilate isomerase/indole-3-glycerol phosphate synthase of Escherichia coli K12 contain five forms of the protein (molecular mass = 49 kDa) which can be separated by isoelectric focusing or two-dimensional gel electrophoresis. The stability of the enzyme and its different forms was studied in exponentially growing, ammonium-starved and energy-depleted cultures using a dual-labeling, pulse-chase method. Labeled enzyme was isolated by standard purification techniques and by immunoprecipitation from crude extracts. The results demonstrated the following: 1) post-translational modification occurred in vivo, 2) modification occurred to the same extent in growing and ammonium-starved cells, 3) modification was dependent on a carbon-energy source for the cells, and 4) the enzyme was not degraded in growing or nongrowing cells. The pulse-chase data also indicated that one form of the protein (band 1) was the precursor of three other forms (bands 3-5) and that the relative amount of one form (band 2) remained constant.