Structure and Mechanism of HpcG, a Hydratase in the Homoprotocatechuate Degradation Pathway of Escherichia coli

Izumi, Atsushi; Rea, Dean; Adachi, Takayuki; Unzai, Satoru; Park, Sam-Yong; Roper, David I.; Tame, J.R.H.

doi:10.1016/j.jmb.2007.05.006

Cited by 19 publications

(46 citation statements)

References 30 publications

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“…Hydrogen bonds and hydrophobic interactions are responsible for a buried surface area of 1,325 Å 2 between these domains. Thus, in contrast to what has been proposed for HpcG (11) , deletion of the NahK NTD might destabilize the incomplete β-barrel and compromise the folding and activity of NahK. As we have observed, residues adjacent to helix H (in the β-barrel) compose the loop which forms the entry of the active site and adopts different conformations upon ligand binding – suggesting that destabilization of this region would decrease (or eliminate) the enzyme activity.…”

Section: Resultscontrasting

confidence: 85%

“…There are also evolutionary questions. Both enzymes belong to the fumarylacetoacetate hydrolase (FAH) superfamily, which includes MhpD from Escherichia coli K-12 (9) , a similar hydratase in the phenylpropionate degradation pathway, and HpcE (10) and HpcG (11) from E. coli C, a decarboxylase and hydratase, respectively, in the homoprotocatechuate degradation pathway. NahK and NahL share 39% sequence identity and likely have the same fold.…”

Section: Introductionmentioning

confidence: 99%

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Crystal Structures of Apo and Liganded 4-Oxalocrotonate Decarboxylase Uncover a Structural Basis for the Metal-Assisted Decarboxylation of a Vinylogous β-Keto Acid

et al. 2016

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The enzymes in the catechol meta-fission pathway have been studied for more than 50 years in several species of bacteria capable of degrading a number of aromatic compounds. In a related pathway, naphthalene, a toxic polycyclic aromatic hydrocarbon, is fully degraded to intermediates of the tricarboxylic acid cycle by the soil bacteria Pseudomonas putida G7. In this organism, the 83 kb NAH7 plasmid carries several genes involved in this biotransformation process. One enzyme in this route, NahK, a 4-oxalocrotonate decarboxylase (4-OD), converts 2-oxo-3-hexenedioate to 2-hydroxy-2,4-pentadienoate using Mg2+ as a cofactor. Efforts to study how 4-OD catalyzes this decarboxylation have been hampered because 4-OD is present in a complex with vinylpyruvate hydratase (VPH), which is the next enzyme in the same pathway. For the first time, a monomeric, stable and active 4-OD has been expressed and purified in the absence of VPH. Crystal structures for NahK in apo form and bonded with five substrate analogues were obtained using two distinct crystallization conditions. Analysis of the crystalline structures implicates a lid domain in substrate binding, and suggests roles for specific residues in a proposed reaction mechanism. In addition, we assign a possible function for the NahK N-terminal domain, which differs from most of the other members of the fumarylacetoacetate hydrolase superfamily. Although the structural basis for metal-dependent β-keto acid decarboxylases has been reported, this is the first structural report for that of a vinylogous β-keto acid decarboxylase and the first crystal structure of a 4-OD.

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Section: Resultscontrasting

confidence: 85%

Section: Introductionmentioning

confidence: 99%

Crystal Structures of Apo and Liganded 4-Oxalocrotonate Decarboxylase Uncover a Structural Basis for the Metal-Assisted Decarboxylation of a Vinylogous β-Keto Acid

et al. 2016

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“…It is possible to fit 936 main-chain atoms of HpcG and MhpD with an rmsd close to 1 Å, omitting small surface loops and disordered regions. 4 The active site of MhpD, inferred from sequence alignment with HpcG, consists of three acidic residues (Glu105, Glu107, and Glu138), which would likely be involved in coordinating the divalent cation essential for activity. The active-site pocket is buried within the core of the FAH fold and situated within the partial β-barrel structure, with the base of the pocket being formed by β-strand 1.…”

Section: Structure Of An Individual Mhpd Subunitmentioning

confidence: 99%

“…The crystal structure of a homologous protein, 2-oxohept-3-ene-1,7-dioate hydratase (HpcG), has been reported recently and solved to a resolution of 1.6 Å (PDB code 2EB4). 4 This enzyme belongs to the homoprotocatechuate degradation pathway of E. coli and also forms a homopentameric assembly. The crystal contains two pentamers stacked together on a common 5-fold axis to form a decamer.…”

Section: Introductionmentioning

confidence: 99%

“…This decamer is also formed in solution, as shown by analytical ultracentrifugation. 4 More remote family members, such as KdhA from Sulfolobus solfataricus 5 and Mus musculus fumarylacetoacetate hydrolase, 6 adopt tetrameric and dimeric forms, respectively. Here we describe the formation of a large dodecahedral protein cage from the E. coli enzyme MhpD.…”

Section: Introductionmentioning

confidence: 99%

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