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Structure of the 2,4'-dihydroxyacetophenone dioxygenase from Alcaligenes sp. 4HAP.

Keegan R, Lebedev A, Erskine P, Guo J, Wood SP, Hopper DJ, Rigby SE, Cooper JB - Acta Crystallogr. D Biol. Crystallogr. (2014)

Bottom Line: The catalytic iron is coordinated by three histidine residues (76, 78 and 114) within a buried active-site cavity.The modelled carbonate is located in a position which is highly likely to be occupied by the α-hydroxyketone group of the bound substrate during catalysis.Modelling of a substrate molecule in this position indicates that it will interact with many conserved amino acids in the predominantly hydrophobic active-site pocket where it undergoes peroxide radical-mediated heterolysis.

View Article: PubMed Central - HTML - PubMed

Affiliation: STFC Rutherford Appleton Laboratory, RAL, Harwell Oxford, Didcot OX11 0FA, England.

ABSTRACT
The enzyme 2,4'-dihydroxyacetophenone dioxygenase (DAD) catalyses the conversion of 2,4'-dihydroxyacetophenone to 4-hydroxybenzoic acid and formic acid with the incorporation of molecular oxygen. Whilst the vast majority of dioxygenases cleave within the aromatic ring of the substrate, DAD is very unusual in that it is involved in C-C bond cleavage in a substituent of the aromatic ring. There is evidence that the enzyme is a homotetramer of 20.3 kDa subunits, each containing nonhaem iron, and its sequence suggests that it belongs to the cupin family of dioxygenases. In this paper, the first X-ray structure of a DAD enzyme from the Gram-negative bacterium Alcaligenes sp. 4HAP is reported, at a resolution of 2.2 Å. The structure establishes that the enzyme adopts a cupin fold, forming dimers with a pronounced hydrophobic interface between the monomers. The catalytic iron is coordinated by three histidine residues (76, 78 and 114) within a buried active-site cavity. The iron also appears to be tightly coordinated by an additional ligand which was putatively assigned as a carbonate dianion since this fits the electron density optimally, although it might also be the product formate. The modelled carbonate is located in a position which is highly likely to be occupied by the α-hydroxyketone group of the bound substrate during catalysis. Modelling of a substrate molecule in this position indicates that it will interact with many conserved amino acids in the predominantly hydrophobic active-site pocket where it undergoes peroxide radical-mediated heterolysis.

No MeSH data available.


Related in: MedlinePlus

The reaction catalysed by 2,4′-dihydroxyacetophenone dioxygenase (DAD). The enzyme has a high affinity for oxygen, which is used for C—C bond cleavage of the substrate (2,4′-dihydroxyacetophenone), yielding 4-hydroxybenzoic acid and formate.
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fig1: The reaction catalysed by 2,4′-dihydroxyacetophenone dioxygenase (DAD). The enzyme has a high affinity for oxygen, which is used for C—C bond cleavage of the substrate (2,4′-dihydroxyacetophenone), yielding 4-hydroxybenzoic acid and formate.

Mentions: The enzyme 2,4′-dihydroxyacetophenone dioxygenase (DAD) catalyses the conversion of 2,4′-dihydroxyaceto­phenone (2,4′-DHAP), a breakdown product of lignin, to 4-hydroxy­benzoic acid and formic acid with the incorporation of molecular oxygen (Fig. 1 ▶). As a bacterial dioxygenase, DAD plays an important environmental role in the aerobic catabolism of aromatic compounds (Gibson & Parales, 2000 ▶; Vaillancourt et al., 2006 ▶). The expression of enzymes such as DAD in appropriately engineered microorganisms has potential for the removal of aromatic pollutants from soil and groundwater and in the production of novel chemicals, for example in biotransformations (Bugg et al., 2011 ▶). Indeed, the bacterial species from which the enzyme originates (Alcaligenes) has been used industrially for biotransformations in the production of unusual amino acids, polyhydroxybutanol and polyhydroxybutyrates for use in new biodegradable plastics (see, for example, Nair et al., 2009 ▶).


Structure of the 2,4'-dihydroxyacetophenone dioxygenase from Alcaligenes sp. 4HAP.

Keegan R, Lebedev A, Erskine P, Guo J, Wood SP, Hopper DJ, Rigby SE, Cooper JB - Acta Crystallogr. D Biol. Crystallogr. (2014)

The reaction catalysed by 2,4′-dihydroxyacetophenone dioxygenase (DAD). The enzyme has a high affinity for oxygen, which is used for C—C bond cleavage of the substrate (2,4′-dihydroxyacetophenone), yielding 4-hydroxybenzoic acid and formate.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4219425&req=5

fig1: The reaction catalysed by 2,4′-dihydroxyacetophenone dioxygenase (DAD). The enzyme has a high affinity for oxygen, which is used for C—C bond cleavage of the substrate (2,4′-dihydroxyacetophenone), yielding 4-hydroxybenzoic acid and formate.
Mentions: The enzyme 2,4′-dihydroxyacetophenone dioxygenase (DAD) catalyses the conversion of 2,4′-dihydroxyaceto­phenone (2,4′-DHAP), a breakdown product of lignin, to 4-hydroxy­benzoic acid and formic acid with the incorporation of molecular oxygen (Fig. 1 ▶). As a bacterial dioxygenase, DAD plays an important environmental role in the aerobic catabolism of aromatic compounds (Gibson & Parales, 2000 ▶; Vaillancourt et al., 2006 ▶). The expression of enzymes such as DAD in appropriately engineered microorganisms has potential for the removal of aromatic pollutants from soil and groundwater and in the production of novel chemicals, for example in biotransformations (Bugg et al., 2011 ▶). Indeed, the bacterial species from which the enzyme originates (Alcaligenes) has been used industrially for biotransformations in the production of unusual amino acids, polyhydroxybutanol and polyhydroxybutyrates for use in new biodegradable plastics (see, for example, Nair et al., 2009 ▶).

Bottom Line: The catalytic iron is coordinated by three histidine residues (76, 78 and 114) within a buried active-site cavity.The modelled carbonate is located in a position which is highly likely to be occupied by the α-hydroxyketone group of the bound substrate during catalysis.Modelling of a substrate molecule in this position indicates that it will interact with many conserved amino acids in the predominantly hydrophobic active-site pocket where it undergoes peroxide radical-mediated heterolysis.

View Article: PubMed Central - HTML - PubMed

Affiliation: STFC Rutherford Appleton Laboratory, RAL, Harwell Oxford, Didcot OX11 0FA, England.

ABSTRACT
The enzyme 2,4'-dihydroxyacetophenone dioxygenase (DAD) catalyses the conversion of 2,4'-dihydroxyacetophenone to 4-hydroxybenzoic acid and formic acid with the incorporation of molecular oxygen. Whilst the vast majority of dioxygenases cleave within the aromatic ring of the substrate, DAD is very unusual in that it is involved in C-C bond cleavage in a substituent of the aromatic ring. There is evidence that the enzyme is a homotetramer of 20.3 kDa subunits, each containing nonhaem iron, and its sequence suggests that it belongs to the cupin family of dioxygenases. In this paper, the first X-ray structure of a DAD enzyme from the Gram-negative bacterium Alcaligenes sp. 4HAP is reported, at a resolution of 2.2 Å. The structure establishes that the enzyme adopts a cupin fold, forming dimers with a pronounced hydrophobic interface between the monomers. The catalytic iron is coordinated by three histidine residues (76, 78 and 114) within a buried active-site cavity. The iron also appears to be tightly coordinated by an additional ligand which was putatively assigned as a carbonate dianion since this fits the electron density optimally, although it might also be the product formate. The modelled carbonate is located in a position which is highly likely to be occupied by the α-hydroxyketone group of the bound substrate during catalysis. Modelling of a substrate molecule in this position indicates that it will interact with many conserved amino acids in the predominantly hydrophobic active-site pocket where it undergoes peroxide radical-mediated heterolysis.

No MeSH data available.


Related in: MedlinePlus