ROLES OF HYDROGEN-BONDING RESIDUES IN THE INTERACTION BETWEEN THE ALPHA-SUBUNITS AND BETA-SUBUNITS IN THE TRYPTOPHAN SYNTHASE COMPLEX - ASN-104 OF THE ALPHA-SUBUNIT IS ESPECIALLY IMPORTANT
K. Hiraga et K. Yutani, ROLES OF HYDROGEN-BONDING RESIDUES IN THE INTERACTION BETWEEN THE ALPHA-SUBUNITS AND BETA-SUBUNITS IN THE TRYPTOPHAN SYNTHASE COMPLEX - ASN-104 OF THE ALPHA-SUBUNIT IS ESPECIALLY IMPORTANT, The Journal of biological chemistry, 272(8), 1997, pp. 4935-4940
The interaction of the a subunit with the beta(2) subunit of tryptopha
n synthase is known to be necessary for the activation of each subunit
and for the catalytic efficiency of the alpha(2) beta(2) complex, To
elucidate the roles of hydrogen bonds in the interaction site between
the alpha and beta subunits for subunit association, eight mutant alph
a subunits at five hydrogen bonding residues (N104D, N104A, N108D, N10
8A, E134A, E135A, N157D, and N157A) were constructed, and the thermody
namic parameters of association with the beta subunit were obtained us
ing a titration calorimeter, The N104D and N104A mutations remarkably
decreased the stimulation activities, the association constants, and t
he association enthalpies, Although the association constant and the s
timulation activities of E134A were reduced in the absence of salt, th
e change in the association enthalpy was relatively small, and the add
ition 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 h
ydrogen bond, The present results suggest that the alpha subunit which
has a mutation at position 104 cannot fold into an intact conformatio
n 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 a
nd mutual activation.