Tyrosine and tryptophan act through the same binding site at the dimer interface of yeast chorismate mutase.
Tyrosine and tryptophan are the regulators of the dimeric yeast chorismate mutase. Biochemical studies reveal two binding sites per molecule for both effectors, tyrosine or tryptophan. A single binding site is built up by helix 8 and helices 4 and 5 of two different subunits. The binding sites have been analyzed in the active enzyme by site directed mutagenesis of critical codons of the coding gene, ARO7. Gly-141 and Ser-142, which both reside on helix 8, are involved in the binding of tyrosine or tryptophan presumably by interacting specifically with the amino- and carboxylate-groups of these amino acid effectors. Interaction with Thr-145 of helix 8 is required for a strong tyrosine binding to the allosteric site. Replacement of Arg-75, which connects helices 4 and 5 or of Arg-76, which is part of helix 5 by alanine residues, resulted in unregulated enzymes. These two residues are bonded to the carboxylate group and phenolic hydroxyl group of tyrosine, respectively, but do not interact with tryptophan by hydrogen bonding in the crystal structures. Phenylalanine, which has low binding affinity slightly activated the chorismate mutase. A T145V mutant chorismate mutase, however, showed increased activation by phenylalanine. Our results support a mechanism by which tyrosine contracts the allosteric site by interacting with its phenolic hydroxyl group. Tryptophan works in an inverse way by opening the allosteric site through the steric size of its side chain.
Schnappauf
G
G
Institut für Mikrobiologie und Genetik, Georg-August-Universität, Grisebachstrasse 8, D-37077 Göttingen, Germany.
Krappmann
S
S
Braus
G H
GH
D000494:Allosteric Regulation; D001665:Binding Sites; D002826:Chorismate Mutase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D003062:Codon; D019281:Dimerization; D005998:Glycine / Q000378:metabolism; D008956:Models, Chemical; D016297:Mutagenesis, Site-Directed; D012441:Saccharomyces cerevisiae / Q000201:enzymology*; D012694:Serine / Q000378:metabolism; D013912:Threonine / Q000378:metabolism; D014364:Tryptophan / Q000378:metabolism*; D014443:Tyrosine / Q000378:metabolism*
D003062:Codon; D013912:Threonine; D014443:Tyrosine; D012694:Serine; D014364:Tryptophan; D002826:Chorismate Mutase; D005998:Glycine