Selective proteolysis of the β2 subunit of Serratia marcescens tryptophan synthase

Abstract

Mild digestion of Serratia marcescens tryptophan synthase β 2 subunit produces a modified β 2 subunit (nicked β 2). The nicked β 2 subunit remains essentially intact and is immunochemically reactive with native β 2 subunit antiserum. Denaturation of the nicked β 2 subunit yields two principal peptide fragments whose minimum molecular weights are 29,500 and 13,400. Loss of enzyme activity is associated with the selective proteolysis. The enzyme cofactor pyridoxal phosphate binding site is on the larger fragment. Following separation of the fragments by urea-gel chromatography, the separated peptides retain immunological cross-reactivity with native β 2 subunit antiserum. These fragments apparently represent two domains that comprise the native Holo β 2 subunit. The immunochemical data suggest that these fragments, when isolated, can assume some tertiary structure and that they may exist as such prior to β monomer or β 2 dimer assembly. The folded fragments may represent intermediates in the biosynthesis of the β 2 subunit as has been suggested for the E. coli enzyme ( A. Högberg-Raibaud and M. E. Goldberg, 1977, Proc. Nat. Acad. Sci. USA 74, 442; Biochemistry 16, 4014).