Active site electronic structure reductase

The initial contribution to this volume provides a detailed overview of how spectroscopy and computations have been used in concert to probe the canonical members of each pyranopterin Mo enzyme family, as well as the pyranopterin dithiolene ligand itself. The discussion focuses on how a combination of enzyme geometric structure, spectroscopy and biochemical data have been used to arrive at an understanding of electronic structure contributions to reactivity in all of the major pyranopterin Mo enzyme families. A unique aspect of this discussion is that spectroscopic studies on relevant small molecule model compounds have been melded with analogous studies on the enzyme systems to arrive at a sophisticated description of active site electronic structure. As the field moves forward, it will become increasingly important to understand the structure, function and reaction mechanisms for the numerous non-canonical [ie. beyond sulfite oxidase, xanthine oxidase, DMSO reductase) pyranopterin Mo enzymes. 

Dinitrogenase has been crystallized and its tertiary structure determined by Kim and Rees. [44, 45,46] As indicated, an Fe-Mo unit serves at the active site. Electrons are furnished to this active unit by the Fe enzyme dinitrogenase reductase. The two units together constitute nitrogenase. 

Clay MD, Yang TC, Jermey FE et al (2006) Geometries and electronic structures of cyanide adducts of the non-heme iron active site of superoxide reductases vibrational and ENDOR studies. Biochemistry 45 427 38... 

Rieske oxygenases are part of a superfamily of enzymes that share a characteristic structure consisting of an oxygenase component (a mononuclear non-heme iron(II) high spin center containing a 2-His-l-carboxylate facial triad motif in the active site) [31-33]. Besides, the active site contains a reductase component (an Fe2-S2 Rieske center) that delivers electrons from NAD(P)H to the oxygenase center [34]. 

Yang YS, Baldwin J, Ley BA, Bollinger Jr JM, Solomon El. 2000. Spectroscopic and electronic structure description of the reduced binuclear non-heme iron active site in ribonucleotide reductase from E. colt comparison to reduced Delta(9) desaturase and electronic structure conttibutions to differences in O2 reactivity. J Am Chem Soc 122 8495-8510. 

Sulfite reductase catalyzes the six-electron reduction by NADPH of sol" to and NO2 to NH3. In E. coli this enzyme is a complex structure with subunit composition 0 8)84 (Siegel et al, 1982). The enzyme active site is on the /3 subunit, which contains both a 4Fe 4S cluster and a siroheme prophyrin. Substrates and ligands have been found to bind to the siroheme. The a subunit binds NADPH and serves to shuttle electrons to the active site through bound FAD and FMN groups. Isolated )8 subunits can catalyze sulfite reduction in the presence of a suitable electron donor. 

As the focus of this review is on copper-dioxygen chemistry, we shall briefly summarize major aspects of the active site chemistry of those proteins involved in 02 processing. The active site structure and chemistry of hemocyanin (He, 02 carrier) and tyrosinase (Tyr, monooxygenase) will be emphasized, since the chemical studies described herein are most relevant to their function. The major classes of these proteins and their origins, primary functions, and leading references are provided in Table 1. Other classes of copper proteins not included here are blue electron carriers [13], copper-thiolate proteins (metallothioneines) [17], and NO reductases (e.g., nitrite [NIR] [18] or nitrous oxide [19]). 

TrxRs are homodimeric flavoproteins [80] that catalyze the NADPH-dependent reduction of thioredoxin (Trx), a ubiquitous 12 kDa protein that is the major protein disulfide reductase in cells [81], and belongs to the pyridine nucleotide-disulfide oxidoreductase family [82]. Each monomer includes an FAD prosthetic group, a NADPH binding site and an active site containing a redox-active selenol group. Electrons are transferred from NADPH via FAD to the active-site selenol of TrxR, which then reduces the substrate Trx [83]. The crystal structure of TrxR is shown in Fig. 13 [84],... 

Valuable spectroscopic studies on the dithiolene chelated to Mo in various enzymes have been enhanced by the knowledge of the structure from X-ray diffraction. Plagued by interference of prosthetic groups—heme, flavin, iron-sulfur clusters—the majority of information has been gleaned from the DMSO reductase system. The spectroscopic tools of X-ray absorption spectroscopy (XAS), electronic ultraviolet/visible (UV/vis) spectroscopy, resonance Raman (RR), MCD, and various electron paramagnetic resonance techniques [EPR, electron spin echo envelope modulation (ESEEM), and electron nuclear double resonance (ENDOR)] have been particularly effective probes of the metal site. Of these, only MCD and RR have detected features attributable to the dithiolene unit. Selected results from a variety of studies are presented below, chosen because their focus is the Mo-dithiolene unit and organized according to method rather than to enzyme or type of active site. 

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