title: Roles of hydrogen bonding residues in the interaction between the alpha and beta subunits in the tryptophan synthase complex. Asn-104 of the alpha subunit is especially important.
abstract: The interaction of the alpha subunit with the beta2 subunit of tryptophan synthase is known to be necessary for the activation of each subunit and for the catalytic efficiency of the alpha2beta2 complex. To elucidate the roles of hydrogen bonds in the interaction site between the alpha and beta subunits for subunit association, eight mutant alpha subunits at five hydrogen bonding residues (N104D, N104A, N108D, N108A, E134A, E135A, N157D, and N157A) were constructed, and the thermodynamic parameters of association with the beta subunit were obtained using a titration calorimeter. The N104D and N104A mutations remarkably decreased the stimulation activities, the association constants, and the association enthalpies. Although the association constant and the stimulation activities of E134A were reduced in the absence of salt, the change in the association enthalpy was relatively small, and the addition of salt could repair its defects. The substitutions at positions 135 and 157 did not affect the stimulation activity and decreased the Gibbs energy of association corresponding to the defect in 1 mol of hydrogen bond. The present results suggest that the alpha subunit which has a mutation at position 104 cannot fold into an intact conformation upon complex formation, resulting in reduced stimulation activities. The hydrogen bond with Asn-104, which is a conserved residue among 16 microorganisms, was especially important for alpha/beta interaction and mutual activation.
authors.affiliation: Institute for Protein Research, Osaka University, 3-2 Yamadaoka, Suita, Osaka 565, Japan.
authors.forename: K
authors.initials: K
authors.lastname: Hiraga
authors.forename: K
authors.initials: K
authors.lastname: Yutani
mesh_terms: D004926:Escherichia coli; D017353:Gene Deletion; D006860:Hydrogen Bonding; D009841:Oligonucleotides; D014367:Tryptophan Synthase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism
chemical_list: D009841:Oligonucleotides; D014367:Tryptophan Synthase