The beta subunit of tryptophan synthase. Clarification of the roles of histidine 86, lysine 87, arginine 148, cysteine 170, and cysteine 230.

摘要: Abstract Our studies, which are aimed at understanding the catalytic mechanism of beta subunit tryptophan synthase from Salmonella typhimurium, use site-directed mutagenesis to clarify functional roles several putative active site residues. Although previous chemical modification studies have suggested that histidine 86, arginine 148, and cysteine 230 essential residues in subunit, our present findings subunits with single amino acid replacements these positions partial activity show 3 not for catalysis or substrate binding. These conclusions consistent recently determined three-dimensional structure alpha 2 complex. Amino substitution lysine 87, forms a Schiff base pyridoxal phosphate wild type yields an inactive form binds phosphate, L-serine. We also report rapid efficient method purifying mutant complex S. typhimurium improved enzyme source. The enzyme, is produced by multicopy plasmid encoding trpA trpB genes expressed Escherichia coli, crystallized crude extracts addition 6% poly(ethylene glycol) 8000 5 mM spermine. This new used accompanying paper purify nine complexes containing subunit.