Importance of Residues 2–9 in the Immunoreactivity, Subunit Interactions, and Activity of the β2 Subunit of Escherichia coli Tryptophan Synthase

The epitope recognized by a monoclonal antibody (mAb19) directed against the beta 2 subunit of Escherichia coli tryptophan synthase was found to be carried by residues 2-9 of the beta chain. The affinities of mAb19 for peptides of different lengths containing the 2-9 sequence were close to 0.6 x 10(9) M-1, the affinity of mAb19 for native beta 2. In view of these results, a model is proposed to account for the kinetics of appearance of the epitope during in vitro renaturation of beta 2 (Murry-Brelier, A., and Goldberg, M.E. (1988) Biochemistry 27, 7633-7640). A mutant producing beta chains lacking residues 1-9 (beta delta 1-9) was prepared. The beta delta 1-9 protein was able to fold into a heat stable homodimer resembling wild type beta 2. Isolated beta delta 1-9 had no detectable enzymatic activity. It could bind alpha chains extremely weakly and be slightly activated. In the presence of the 1-9 peptide, the beta delta 1-9 protein could bind alpha chains much more strongly and generate a 50% active enzyme. Thus, although having little role in the overall folding and stability of the protein, the 1-9 sequence of the beta chain appears strongly involved in the alpha-beta interactions and in the enzymatic activity.