Effect of amino acid residues on conformational stability in eight mutant proteins variously substituted at a unique position of the tryptophan synthase alpha-subunit.
To elucidate the role of individual amino acid residues in stabilizing the conformation of a protein, the stabilities of wild-type tryptophan synthase alpha-subunit from Escherichia coli and seven mutant proteins substituted by single amino acid residues at position 49, which is buried in the interior of the protein, were compared. The mutant proteins have Gln, Met, Val, Tyr, Leu, Ser, or Lys in place of Glu in the wild-type protein. The dissociation constant, pK, of the Glu residue at position 49 for the wild-type protein was determined to be 7.5 from a titration curve obtained by comparison of two-dimensional isoelectric focusing electrophoresis of the wild-type and mutant proteins. Our results indicate that 1) the conformational stabilities of the proteins studied increase linearly with hydrophobicity of the substituting residues (except Tyr), with the coefficient of this linear dependence being 2.0, 3.4, or 1.3 at pH 5.5, 7.0, or 9.0, respectively; and 2) Lys or Glu at position 49 serve as a destabilizing factor when ionized.
D000596:Amino Acids / Q000032:analysis; D004926:Escherichia coli / Q000201:enzymology; D006863:Hydrogen-Ion Concentration; D007525:Isoelectric Focusing; D009154:Mutation; D011487:Protein Conformation; D013329:Structure-Activity Relationship; D014367:Tryptophan Synthase / Q000032:analysis* / Q000235:genetics
D000596:Amino Acids; D014367:Tryptophan Synthase