A Bacterial Tyrosine Aminomutase Proceeds through Retention or Inversion of Stereochemistry To Catalyze Its Isomerization Reaction

Abstract

β-Amino acids are biologically active compounds of interest in medicinal chemistry. A class I lyase-like family of aminomutases isomerizes (S)-α-arylalanines to the corresponding β-amino acids by exchange of the NH2/H pair. This family uses a 3,5-dihydro-5-methylidene-4H-imidazol-4-one (MIO) group within the active site to initiate the reaction. The absolute stereochemistry of the product is known for an MIO-dependent tyrosine aminomutase from Chondromyces crocatus (CcTAM) that isomerizes (S)-α-tyrosine to (R)-β-tyrosine. To evaluate the cryptic stereochemistry of the CcTAM mechanism, (2S,3S)-[2,3-(2)H2]- and (2S,3R)-[3-(2)H]-α-tyrosine were stereoselectively synthesized from unlabeled (or [(2)H]-labeled) (4'-hydroxyphenyl)acrylic acids by reduction with D2 (or H2) gas and a chiral Rh-Prophos catalyst. GC/EIMS analysis of the [(2)H]-β-tyrosine biosynthesized by CcTAM revealed that the α-amino group was transferred to Cβ of the phenylpropanoid skeleton with retention of configuration. These labeled substrates also showed that the pro-(3S) proton exchanges with protons from the bulk media during its migration to Cα during catalysis. (1)H- and (2)H NMR analyses of the [(2)H]-β-tyrosine derived from (2S)-[3,3-(2)H2]-α-tyrosine by CcTAM catalysis showed that the migratory proton attached to Cα of the product also with retention of configuration. CcTAM is stereoselective for (R)-β-tyrosine (85%) yet also forms the (S)-β-tyrosine enantiomer (15%) through inversion of configuration at both migration termini, as described herein. The proportion of the (S)-β-isomer made by CcTAM during steady state interestingly increased with solvent pH, and this effect on the proposed reaction mechanism is also discussed.