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PDBsum entry 2vss
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References listed in PDB file
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Key reference
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Title
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A ternary complex of hydroxycinnamoyl-Coa hydratase-Lyase (hchl) with acetyl-Coa and vanillin gives insights into substrate specificity and mechanism.
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Authors
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J.P.Bennett,
L.Bertin,
B.Moulton,
I.J.Fairlamb,
A.M.Brzozowski,
N.J.Walton,
G.Grogan.
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Ref.
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Biochem J, 2008,
414,
281-289.
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PubMed id
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Abstract
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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.
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