4X2P

P. putida mandelate racemase in complex with 3-hydroxypyruvate

Summary for 4X2P

• Entry DOI: 10.2210/pdb4x2p/pdb
• Descriptor: Mandelate racemase, 3-HYDROXYPYRUVIC ACID, MAGNESIUM ION, ... (4 entities in total)
• Functional Keywords: racemase, enolase superfamily, isomerase
• Biological source: Pseudomonas putida
• Total number of polymer chains: 1
• Total formula weight: 41432.99

Authors

Wyatt, B.N.,St.Maurice, M. (deposition date: 2014-11-26, release date: 2015-10-14, Last modification date: 2024-11-20)

Primary citation

Nagar, M.,Wyatt, B.N.,St.Maurice, M.,Bearne, S.L.
Inactivation of Mandelate Racemase by 3-Hydroxypyruvate Reveals a Potential Mechanistic Link between Enzyme Superfamilies.
Biochemistry, 54:2747-2757, 2015

PubMed Abstract: Mandelate racemase (MR), a member of the enolase superfamily, catalyzes the Mg(2+)-dependent interconversion of the enantiomers of mandelate. Several α-keto acids are modest competitive inhibitors of MR [e.g., mesoxalate (Ki = 1.8 ± 0.3 mM) and 3-fluoropyruvate (Ki = 1.3 ± 0.1 mM)], but, surprisingly, 3-hydroxypyruvate (3-HP) is an irreversible, time-dependent inhibitor (kinact/KI = 83 ± 8 M(-1) s(-1)). Protection from inactivation by the competitive inhibitor benzohydroxamate, trypsinolysis and electrospray ionization tandem mass spectrometry analyses, and X-ray crystallographic studies reveal that 3-HP undergoes Schiff-base formation with Lys 166 at the active site, followed by formation of an aldehyde/enol(ate) adduct. Such a reaction is unprecedented in the enolase superfamily and may be a relic of an activity possessed by a promiscuous progenitor enzyme. The ability of MR to form and deprotonate a Schiff-base intermediate furnishes a previously unrecognized mechanistic link to other α/β-barrel enzymes utilizing Schiff-base chemistry and is in accord with the sequence- and structure-based hypothesis that members of the metal-dependent enolase superfamily and the Schiff-base-forming N-acetylneuraminate lyase superfamily and aldolases share a common ancestor.

PubMed: 25844917
DOI: 10.1021/acs.biochem.5b00221

Experimental method

X-RAY DIFFRACTION (1.65 Å)