Pyranopterin conformation defines the function of molybdenum and tungsten enzymes

Abstract

We have analyzed the conformations of 319 pyranopterins in 102 protein structures of mononuclear molybdenum and tungsten enzymes. These span a continuum between geometries anticipated for quinonoid dihydro, tetrahydro, and dihydro oxidation states. We demonstrate that pyranopterin conformation is correlated with the protein folds defining the three major mononuclear molybdenum and tungsten enzyme families, and that binding-site microtuning controls pyranopterin oxidation state. Enzymes belonging to the bacterial dimethyl sulfoxide reductase (DMSOR) family contain a metal-bis-pyranopterin cofactor, the two pyranopterins of which have distinct conformations, with one similar to the predicted tetrahydro form, and the other similar to the predicted dihydro form. Enzymes containing a single pyranopterin belong to either the xanthine dehydrogenase (XDH) or sulfite oxidase (SUOX) families, and these have pyranopterin conformations similar to those predicted for tetrahydro and dihydro forms, respectively. This work provides keen insight into the roles of pyranopterin conformation and oxidation state in catalysis, redox potential modulation of the metal site, and catalytic function.energetics | molybdoenzymes | redox chemistry T he pyranopterin dithiolene ligand is present in all molybdenum (Mo) and tungsten (W) containing enzymes with the exception of nitrogenase (1). These enzymes, known as mononuclear Mo/W enzymes, play pivotal roles in metabolism, global geochemical cycles, and microbial metabolic diversity (2-5). Their active sites, comprising a Mo or W ion and one or two pyranopterins, catalyze a diversity of redox transformations spanning a reduction potential range of approximately one volt. While the immediate environment of the substrate-binding metal ion is critically important in catalysis (6), little attention has been focused on the impact of variations in pyranopterin structure on enzyme function.Protein-bound pyranopterins are typically interpreted as having the tricyclic structure depicted in Fig. 1A, comprising pyrimidine, piperazine, and pyran-dithiolene rings. The dithiolene chelate binds a single Mo/W atom, which constitutes the catalytic active site. The pyranopterin shown in Fig. 1A is also known as the tetrahydro mononucleotide form due to the oxidation state of its piperazine ring and the presence of a phosphomethyl group attached to its C-2 atom (1). This form is assigned to the xanthine dehydrogenases, and in its cytosine dinucleotide form to the bacterial carbon monoxide dehydrogenases and aldehyde dehydrogenases (these enzymes are referred to collectively as the XDH family). It is also assigned to the sulfite oxidases and plant nitrate reductases [SUOX family (1, 5)].Mononuclear Mo/W enzymes also coordinate metal-bis-pyranopterin cofactors, either as the metal-bis(pyranopterin guanine dinucleotide) form (Fig. 1D) found in the dimethyl sulfoxide reductase family of molybdoenzyme subunits (DMSOR family), or the mononuclear bis-pyranopterin form found in the thermophilic aldehyde oxidoreductases...

Electrochemical evidence that pyranopterin redox chemistry controls the catalysis of YedY, a mononuclear Mo enzyme

“…A 2012 computational study provided evidence that two different oxidation states can be accessed by protein-bound pyranopterin ligands (10). Conformational analysis and electronic structure calculations were used to assign redox states to the pyranopterin ligands in all known mononuclear Mo enzyme structures (10).…”

Assembly pathway of a bacterial complex iron sulfur molybdoenzyme

“…With DmsAB assembled, it is targeted to the Tat translocon (9-11) and translocated (12)(13)(14). Finally, DmsAB docks to the membrane anchor subunit DmsC to complete the accomplishment of securing a fully functional respiratory enzyme (15).…”

“…This pathway is illustrated in Figure 2. The cartooned pathway shows stages (10)(11)(12)(13)(14)(15). The nascent chain exiting the ribosome and the RR-leader interacts with DnaK and trigger factor (TF) (1 and 2).…”

“…DMSO reductase is an archetypical heterotrimer CISM enzyme comprised of three modular subunits, DmsABC (15,39). DmsA, the RR-leader containing subunit of the enzyme, functions as the catalytic subunit that coordinates the MobisPGD catalytic cofactor in addition to one [4Fe-4S] (40,41).…”

“…Respiratory oxidoreductases that contain a molybdenum-bis(pyranopterin guanine dinucleotide) (Mo-bisPGD) catalytic cofactor are grouped under the molybdoenzyme superfamily (13)(14)(15)(16)(17). In many instances, the catalytic subunit also coordinates an iron-sulfur cluster ([Fe-S]) near the catalytic site, an electron transfer subunit that typically contains typically four [Fe-S] clusters of various Fe:S ratios, and a membrane anchor subunit required to connect the [Fe-S] cluster electrical conduit to the quinone pool (16,18) which may also contain hemes.…”