2VSU

A ternary complex of Hydroxycinnamoyl-CoA Hydratase-Lyase (HCHL) with acetyl-Coenzyme A and vanillin gives insights into substrate specificity and mechanism.

Summary for 2VSU

• Entry DOI: 10.2210/pdb2vsu/pdb
• Related: 2J5I 2VSS
• Descriptor: P-HYDROXYCINNAMOYL COA HYDRATASE/LYASE, ACETYL COENZYME *A, 4-hydroxy-3-methoxybenzaldehyde, ... (7 entities in total)
• Functional Keywords: lyase, aldolase, crotonase, hydratase
• Biological source: PSEUDOMONAS FLUORESCENS; More
• Total number of polymer chains: 6
• Total formula weight: 190247.55

Authors

Bennett, J.P.,Bertin, L.M.,Brzozowski, A.M.,Walton, N.J.,Grogan, G. (deposition date: 2008-04-29, release date: 2008-05-27, Last modification date: 2023-12-13)

Primary citation

Bennett, J.P.,Bertin, L.,Moulton, B.,Fairlamb, I.J.S.,Brzozowski, A.M.,Walton, N.J.,Grogan, G.
A Ternary Complex of Hydroxycinnamoyl-Coa Hydratase-Lyase (Hchl) with Acetyl-Coa and Vanillin Gives Insights Into Substrate Specificity and Mechanism.
Biochem.J., 414:281-, 2008

PubMed Abstract: HCHL (hydroxycinnamoyl-CoA hydratase-lyase) catalyses the biotransformation of feruloyl-CoA to acetyl-CoA and the important flavour-fragrance compound vanillin (4-hydroxy-3-methoxybenzaldehyde) and is exploited in whole-cell systems for the bioconversion of ferulic acid into natural equivalent vanillin. The reaction catalysed by HCHL has been thought to proceed by a two-step process involving first the hydration of the double bond of feruloyl-CoA and then the cleavage of the resultant beta-hydroxy thioester by retro-aldol reaction to yield the products. Kinetic analysis of active-site residues identified using the crystal structure of HCHL revealed that while Glu-143 was essential for activity, Ser-123 played no major role in catalysis. However, mutation of Tyr-239 to Phe greatly increased the K(M) for the substrate ferulic acid, fulfilling its anticipated role as a factor in substrate binding. Structures of WT (wild-type) HCHL and of the S123A mutant, each of which had been co-crystallized with feruloyl-CoA, reveal a subtle helix movement upon ligand binding, the consequence of which is to bring the phenolic hydroxyl of Tyr-239 into close proximity to Tyr-75 from a neighbouring subunit in order to bind the phenolic hydroxyl of the product vanillin, for which electron density was observed. The active-site residues of ligand-bound HCHL display a remarkable three-dimensional overlap with those of a structurally unrelated enzyme, vanillyl alcohol oxidase, that also recognizes p-hydroxylated aromatic substrates related to vanillin. The data both explain the observed substrate specificity of HCHL for p-hydroxylated cinnamate derivatives and illustrate a remarkable convergence of the molecular determinants of ligand recognition between the two otherwise unrelated enzymes.

PubMed: 18479250
DOI: 10.1042/BJ20080714

Experimental method

X-RAY DIFFRACTION (1.9 Å)