Abstract
Chorismate synthase catalyzes the last step in the common shikimate pathway leading to aromatic compounds such as the aromatic amino acids. The reaction consists of the 1,4-anti-elimination of the 3-phosphate group and the C-(6proR) hydrogen from 5-enolpyruvylshikimate 3-phosphate to yield chorismate. Although this reaction does not involve a net redox change, the enzyme has an absolute requirement for reduced flavin mononucleotide, which is not consumed during the reaction. Two invariant histidine residues are found in the active site of the enzyme: His(17) and His(106). Using site-directed mutagenesis, both histidines were replaced by alanine, reducing the activity 10- and 20-fold in the H106A and H17A mutant protein, respectively. Based on the characterization of the two single mutant proteins, it is proposed that His(106) serves to protonate the monoanionic reduced FMN, whereas His(17) protonates the leaving phosphate group of the substrate. An enzymatic reaction mechanism in keeping with the experimental results is presented.
Highlights
Chorismate synthase catalyzes the last step in the common shikimate pathway leading to aromatic compounds such as the aromatic amino acids
This reaction does not involve a net redox change, the enzyme has an absolute requirement for reduced flavin mononucleotide, which is not consumed during the reaction
Chorismate synthase catalyzes the last step in the shikimate pathway leading to the branch point metabolite chorismate, which is utilized in a number of enzymatic transformations toward the biosynthesis of aromatic compounds such as the aromatic amino acids tyrosine, phenylalanine, and tryptophan
Summary
Chorismate synthase catalyzes the last step in the common shikimate pathway leading to aromatic compounds such as the aromatic amino acids. Chorismate synthase catalyzes the last step in the shikimate pathway leading to the branch point metabolite chorismate, which is utilized in a number of enzymatic transformations toward the biosynthesis of aromatic compounds such as the aromatic amino acids tyrosine, phenylalanine, and tryptophan (for a recent review, see Ref. 1) This reaction involves an 1,4-anti-elimination of the 3-phosphate group and the C-(6proR) hydrogen from 5-enolpyruvylshikimate-3-phosphate (EPSP)1 [2, 3]. In the reported structure of the ternary complex, comprising FMN and EPSP as ligands, EPSP is stacked above the si-face of the isoalloxazine ring in an average distance of 3.3 Å This close juxtaposition of FMN and EPSP supports earlier suggestions for a direct role of reduced FMN in the elimination reaction, such as a radical mechanism, in which electron transfer from the reduced cofactor to the substrate initiates C–O bond breakage of the phosphate group to yield a substrate-derived neutral radical (6 –9, 11). In contrast to the mode of flavin reduction, the two classes of chorismate synthases show no apparent difference in the mechanism of the elimination reaction
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