title: Protein-protein interactions in the β-oxidation part of the phenylacetate utilization pathway: crystal structure of the PaaF-PaaG hydratase-isomerase complex.
abstract: Microbial anaerobic and so-called hybrid pathways for degradation of aromatic compounds contain β-oxidation-like steps. These reactions convert the product of the opening of the aromatic ring to common metabolites. The hybrid phenylacetate degradation pathway is encoded in Escherichia coli by the paa operon containing genes for 10 enzymes. Previously, we have analyzed protein-protein interactions among the enzymes catalyzing the initial oxidation steps in the paa pathway (Grishin, A. M., Ajamian, E., Tao, L., Zhang, L., Menard, R., and Cygler, M. (2011) J. Biol. Chem. 286, 10735-10743). Here we report characterization of interactions between the remaining enzymes of this pathway and show another stable complex, PaaFG, an enoyl-CoA hydratase and enoyl-Coa isomerase, both belonging to the crotonase superfamily. These steps are biochemically similar to the well studied fatty acid β-oxidation, which can be catalyzed by individual monofunctional enzymes, multifunctional enzymes comprising several domains, or enzymatic complexes such as the bacterial fatty acid β-oxidation complex. We have determined the structure of the PaaFG complex and determined that although individually PaaF and PaaG are similar to enzymes from the fatty acid β-oxidation pathway, the structure of the complex is dissimilar from bacterial fatty acid β-oxidation complexes. The PaaFG complex has a four-layered structure composed of homotrimeric discs of PaaF and PaaG. The active sites of PaaF and PaaG are adapted to accept the intermediary components of the Paa pathway, different from those of the fatty acid β-oxidation. The association of PaaF and PaaG into a stable complex might serve to speed up the steps of the pathway following the conversion of phenylacetyl-CoA to a toxic and unstable epoxide-CoA by PaaABCE monooxygenase.
authors.affiliation: Department of Biochemistry, University of Saskatchewan, Saskatoon, Saskatchewan S7N 5E5, Canada.
authors.forename: Andrey M
authors.initials: AM
authors.lastname: Grishin
authors.forename: Eunice
authors.initials: E
authors.lastname: Ajamian
authors.forename: Linhua
authors.initials: L
authors.lastname: Zhang
authors.forename: Isabelle
authors.initials: I
authors.lastname: Rouiller
authors.forename: Mihnea
authors.initials: M
authors.lastname: Bostina
authors.forename: Miroslaw
authors.initials: M
authors.lastname: Cygler
mesh_terms: D000105:Acetyl Coenzyme A / Q000737:chemistry / Q000378:metabolism; D019748:Carbon-Carbon Double Bond Isomerases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D020134:Catalytic Domain; D018360:Crystallography, X-Ray; D064006:Dodecenoyl-CoA Isomerase; D004746:Enoyl-CoA Hydratase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics / Q000378:metabolism; D029968:Escherichia coli Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D008854:Microscopy, Electron; D008956:Models, Chemical; D008958:Models, Molecular; D015394:Molecular Structure; D046912:Multiprotein Complexes / Q000737:chemistry / Q000378:metabolism / Q000648:ultrastructure; D009876:Operon / Q000235:genetics; D010084:Oxidation-Reduction; D010648:Phenylacetates / Q000737:chemistry* / Q000378:metabolism; D011485:Protein Binding; D055503:Protein Multimerization; D020836:Protein Structure, Quaternary; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D014867:Water / Q000737:chemistry / Q000378:metabolism
chemical_list: D029968:Escherichia coli Proteins; D046912:Multiprotein Complexes; D010648:Phenylacetates; D014867:Water; D000105:Acetyl Coenzyme A; C037816:phenylacetyl-coenzyme A; D004746:Enoyl-CoA Hydratase; C577533:PaaF protein, E coli; C577534:PaaG protein, E coli; D019748:Carbon-Carbon Double Bond Isomerases; D064006:Dodecenoyl-CoA Isomerase