PDBsum entry 3f6b

Lyase

PDB id: 3f6b

Contents

Protein chain

525 a.a. *

Ligands

8PA

Metals

_MG

Waters ×462

* Residue conservation analysis

PDB id:

3f6b

Name:

Lyase

Title:

Crystal structure of benzoylformate decarboxylase in complex with the pyridyl inhibitor paa

Structure:

Benzoylformate decarboxylase. Chain: x. Synonym: bfd, bfdc. Engineered: yes

Source:

Pseudomonas putida. Organism_taxid: 303. Strain: dsm 291 / ncib 9494 / nctc 10936 / stanier 90. Atcc: 12633. Gene: mdlc. Expressed in: escherichia coli. Expression_system_taxid: 562.

Resolution:

1.34Å R-factor: 0.163 R-free: 0.182

Authors:

G.S.Brandt,M.J.Mcleish,G.L.Kenyon,G.A.Petsko,D.Ringe,F.Jordan

Key ref:

S.Chakraborty et al. (2009). Detection and time course of formation of major thiamin diphosphate-bound covalent intermediates derived from a chromophoric substrate analogue on benzoylformate decarboxylase. Biochemistry, 48, 981-994. PubMed id: 19140682

Date:

05-Nov-08 Release date: 09-Dec-08

Protein chain

P20906  (MDLC_PSEPU) -  Benzoylformate decarboxylase from Pseudomonas putida

Seq: 528 a.a.

Struc: 525 a.a.

Enzyme reactions

• Enzyme class: E.C.4.1.1.7 - benzoylformate decarboxylase.
• Reaction: phenylglyoxylate + H⁺ = benzaldehyde + CO2
• Cofactor: Thiamine diphosphate

Thiamine diphosphate (Bound ligand (Het Group name = 8PA) matches with 72.22% similarity)

Molecule diagrams generated from .mol files obtained from the KEGG ftp site

reference

Biochemistry 48:981-994 (2009)

PubMed id: 19140682

Detection and time course of formation of major thiamin diphosphate-bound covalent intermediates derived from a chromophoric substrate analogue on benzoylformate decarboxylase.

S.Chakraborty, N.S.Nemeria, A.Balakrishnan, G.S.Brandt, M.M.Kneen, A.Yep, M.J.McLeish, G.L.Kenyon, G.A.Petsko, D.Ringe, F.Jordan.

ABSTRACT

The mechanism of the enzyme benzoylformate decarboxylase (BFDC), which carries out a typical thiamin diphosphate (ThDP)-dependent nonoxidative decarboxylation reaction, was studied with the chromophoric alternate substrate (E)-2-oxo-4(pyridin-3-yl)-3-butenoic acid (3-PKB). Addition of 3-PKB resulted in the appearance of two transient intermediates formed consecutively, the first one to be formed a predecarboxylation ThDP-bound intermediate with lambda(max) at 477 nm, and the second one corresponding to the first postdecarboxylation intermediate the enamine with lambda(max) at 437 nm. The time course of formation/depletion of the PKB-ThDP covalent complex and of the enamine showed that decarboxylation was slower than formation of the PKB-ThDP covalent adduct. When the product of decarboxylation 3-(pyridin-3-yl)acrylaldehyde (PAA) was added to BFDC, again an absorbance with lambda(max) at 473 nm was formed, corresponding to the tetrahedral adduct of PAA with ThDP. Addition of well-formed crystals of BFDC to a solution of PAA resulted in a high resolution (1.34 A) structure of the BFDC-bound adduct of ThDP with PAA confirming the tetrahedral nature at the C2alpha atom, rather than of the enamine, and supporting the assignment of the lambda(max) at 473 nm to the PAA-ThDP adduct. The structure of the PAA-ThDP covalent complex is the first example of a product-ThDP adduct on BFDC. Similar studies with 3-PKB indicated that decarboxylation had taken place. Evidence was also obtained for the slow formation of the enamine intermediate when BFDC was incubated with benzaldehyde, the product of the decarboxylation reaction thus confirming its presence on the reaction pathway.