A Paradigm for CH Bond Cleavage: Structural and Functional Aspects of Transition State Stabilization by Mandelate Racemase

Document Type

Conference Proceeding

Language

eng

Publication Date

6-2017

Publisher

Elsevier

Source Publication

Advances in Protein Chemistry and Structural Biology

Source ISSN

9780128118764

Abstract

Mandelate racemase (MR) from Pseudomonas putida catalyzes the Mg2+-dependent, 1,1-proton transfer reaction that racemizes (R)- and (S)-mandelate. MR shares a partial reaction (i.e., the metal ion-assisted, Brønsted base-catalyzed proton abstraction of the α-proton of carboxylic acid substrates) and structural features ((β/α)7β-barrel and N-terminal α + β capping domains) with a vast group of homologous, yet functionally diverse, enzymes in the enolase superfamily. Mechanistic and structural studies have developed this enzyme into a paradigm for understanding how enzymes such as those of the enolase superfamily overcome kinetic and thermodynamic barriers to catalyze the abstraction of an α-proton from a carbon acid substrate with a relatively high pKa value. Structural studies on MR bound to intermediate/transition state analogues have delineated those structural features that MR uses to stabilize transition states and enhance reaction rates of proton abstraction. Kinetic, site-directed mutagenesis, and structural studies have also revealed that the phenyl ring of the substrate migrates through the hydrophobic cavity within the active site during catalysis and that the Brønsted acid–base catalysts (Lys 166 and His 297) may be utilized as binding determinants for inhibitor recognition. In addition, structural studies on the adduct formed from the irreversible inhibition of MR by 3-hydroxypyruvate revealed that MR can form and deprotonate a Schiff-base with 3-hydroxypyruvate to yield an enol(ate)-aldehyde adduct, suggesting a possible evolutionary link between MR and the Schiff-base forming aldolases. As the archetype of the enolase superfamily, mechanistic and structural studies on MR will continue to enhance our understanding of enzyme catalysis and furnish insights into the evolution of enzyme function.

Comments

Advances in Protein Chemistry and Structural Biology, Vol. 109 (2017): 113-160. DOI.

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