4BK3

Crystal structure of 3-hydroxybenzoate 6-hydroxylase uncovers lipid- assisted flavoprotein strategy for regioselective aromatic hydroxylation: Y105F mutant

Summary for 4BK3

• Entry DOI: 10.2210/pdb4bk3/pdb
• Related: 4BJY 4BJZ 4BK1 4BK2
• Descriptor: PROBABLE SALICYLATE MONOOXYGENASE, FLAVIN-ADENINE DINUCLEOTIDE, PHOSPHATIDYLGLYCEROL-PHOSPHOGLYCEROL, ... (5 entities in total)
• Functional Keywords: oxidoreductase, flavoprotein, gentisate, hydroxylase, monooxygenase, phospholipid
• Biological source: RHODOCOCCUS JOSTII
• Total number of polymer chains: 1
• Total formula weight: 48661.70

Authors

Orru, R.,Montersino, S.,Barendregt, A.,Westphal, A.H.,van Duijn, E.,Mattevi, A.,van Berkel, W.J.H. (deposition date: 2013-04-21, release date: 2013-07-24, Last modification date: 2023-12-20)

Primary citation

Montersino, S.,Orru, R.,Barendregt, A.,Westphal, A.H.,Van Duijn, E.,Mattevi, A.,Van Berkel, W.J.H.
Crystal Structure of 3-Hydroxybenzoate 6-Hydroxylase Uncovers Lipid-Assisted Flavoprotein Strategy for Regioselective Aromatic Hydroxylation
J.Biol.Chem., 288:26235-, 2013

PubMed Abstract: 3-Hydroxybenzoate 6-hydroxylase (3HB6H) from Rhodococcus jostii RHA1 is a dimeric flavoprotein that catalyzes the NADH- and oxygen-dependent para-hydroxylation of 3-hydroxybenzoate to 2,5-dihydroxybenzoate. In this study, we report the crystal structure of 3HB6H as expressed in Escherichia coli. The overall fold of 3HB6H is similar to that of p-hydroxybenzoate hydroxylase and other flavoprotein aromatic hydroxylases. Unexpectedly, a lipid ligand is bound to each 3HB6H monomer. Mass spectral analysis identified the ligand as a mixture of phosphatidylglycerol and phosphatidylethanolamine. The fatty acid chains occupy hydrophobic channels that deeply penetrate into the interior of the substrate-binding domain of each subunit, whereas the hydrophilic part is exposed on the protein surface, connecting the dimerization domains via a few interactions. Most remarkably, the terminal part of a phospholipid acyl chain is directly involved in the substrate-binding site. Co-crystallized chloride ion and the crystal structure of the H213S variant with bound 3-hydroxybenzoate provide hints about oxygen activation and substrate hydroxylation. Essential roles are played by His-213 in catalysis and Tyr-105 in substrate binding. This phospholipid-assisted strategy to control regioselective aromatic hydroxylation is of relevance for optimization of flavin-dependent biocatalysts.

PubMed: 23864660
DOI: 10.1074/JBC.M113.479303

Experimental method

X-RAY DIFFRACTION (1.78 Å)