Spectroscopic studies reductase

Caldeira J, R Feicht, H White, M Teixeira, JJG Moura, H Simon, I Moura (1996) EPR and Mdssbauer spectroscopic studies on enoate reductase. J Biol Chem 271 18743-18748. 

Gomes CM, Vicente JB,Wasserfallen A,Teixeira M. 2000. Spectroscopic studies and characterization of a novel electron-transfer chain from Escherichia coli involving a flavorubredoxin and its flavoprotein reductase partner. Biochemistry 39 16230-7. 

Spectroscopic studies indicate that the siroheme and 4Fe 4S cluster are exchange coupled in all oxidation states (Christner et al., 1984 Cline et al, 1985a,b). The structural basis of this coupling has been provided by a preliminary 3 A resolution X-ray structure of sulfite reductase in the oxidized state (McRee et al., 1986). The siroheme and the 4Fe 4S cluster are packed against each other and appear to share a common ligand (Fig. 18). The distance from the siroheme iron to the cluster center is... 

Three aldose reductase inhibitors (490 492) were isolated from Dictyodendrilla sp. from Japan and their structures were determined by X-ray analysis and spectroscopic studies [418],... 

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. 

One example of an enzyme-catalyzed reaction involving a radical intermediate is the enzyme ribonucleotide reductase, which catalyzes the conversion of ribonucleotides (used for RNA biosynthesis) to 2 -deoxyribonucleotides (used for DNA biosynthesis), as illustrated in Fig. 16. Spectroscopic studies of the R2 subunit of Escherichia coli ribonucleotide reductase have shown that it can form a stable, long-lived, tyrosyl radical species—the first protein radical to be discovered (13). 

Anderson, L.J., Protein film voltammetry and spectroscopic studies ofibacterial nitrate reductases, Ph.D. Thesis, University of East Anglia, Norwich, U.K, 2002. 

Lampreia J, Moura L, Teixeria M, Peck HD Jr, LeGall J, Huynth BH, Moura JJG (1990) The active centers of adenylylsulfate reductase from Desulfovibrio gigas. Characterization and spectroscopic studies. Eur J Biochem 188 653-664... 

Spectroscopic studies indicate homolytic cleavage of the enzyme bound coenzyme to a deoxyadenosyl radical (ACH2 ) and Bi2r as a common characteristic of these enzymatic reactions. Isomerization, described above, involves an apparent intramolecular 1,2-shift of a hydrogen and an electronegative group (X = OH, NH2, Table 1, entry 4 to 6) or a carbon skeleton (Table 1, entry 7 to 10). However, in the nucleotide reductase system, coenzyme B12 has a unique role of radical initiator in a radical chain mechanism rather... 

The terminal step in methane generation by several methanogenic organisms, of which the best studied is the archaeon Methanobacterium thermoautotrophicum, is catalyzed by the enzyme S-methyl coenzyme M reductase (methylreductase, EC 1.8.-.-). This enzyme contains a macrocyclic tetrapyrrole-derived cofactor, F430, at the active site coordinating Ni(II) in the resting state. A Ni(I) state (Ni1F430) has been proposed as the active form of the cofactor. Extensive mechanistic and spectroscopic studies have been performed on the holoenzyme, isolated cofactor, and various synthetic model compounds. These studies are summarized in... 

Spectroscopic studies of Pyrococcus furiosus superoxide reductase Implications for active-site structures and the catalytic mechanism. J. Am. Chem. Soc. 124, 788-805. 

The use of a mixed-valent, dinuclear iron site, similar to those in hemerythrin and ribonucleotide reductase,to catalyze a nonredox reaction such as phosphate ester hydrolysis is novel and unexpected for a variant of the familiar oxo(hydroxo)-bridged diiron center. In contrast to the general agreement that exists regarding the spectroscopic and physical properties of the PAPs, their kinetics properties and especially their mechanism of action remain controversial. Much of the disagreement stems from the different pH dependences of the catalytic activity of BSPAP and Uf, which is due to the fact that the former is isolated in a proteolytically activated form while the latter is not. Proteolysis results in a substantial increase in optimal pH in addition to an increase in catalytic activity at the optimal pH. "" Current data suggest that many of the spectroscopic studies described in the literature were performed on a catalytically inactive form of the enzyme. As a result, the roles of the trivalent and divalent metal ions in catalysis and in particular the identity of the nucleophilic hydroxide that directly attacks the phosphate ester remain unresolved. 

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. 

Hagedoorn et al. have investigated the redox properties of tungsten-substituted DMSO reductase from R. capsulatus The enzyme is produced by growing R. capsulatus cells on tungstate rather than molybdate and structural and spectroscopic studies have shown that the W-enzyme exhibits similar features to its native Mo analogue. An EPR redox titration concentration of the W state determined and redox potentials of -194 mV and -134 mV vs. NHE, respectively, at pH 7.0, which are considerably lower than the values reported for the Mo-containing R. sphaeroides DMSO reductase. These redox potentials are consistent with the known ability of (Mo) DMSO reductase to catalyze both the reduction of DMSO and the oxidation of DMS. 

Christner, J.A., E. Munck, P.A. Janick, and L.M. Siegel (1981). Mossbauer spectroscopic studies of Escherichia coli sulfite reductase. J. Biol. Chem. 256, 2098-2101. 

Wei PP, Skulan AJ, Mitic N, Yang YS, Saleh L, Bollinger Jr JM, Solomon El. 2004. Electronic and spectroscopic studies of the non-heme reduced binuclear iron sites of two ribonucleotide reductase variants comparison to reduced methane monooxygenase and contributions to O2 reactivity. JAm Chem Soc 126 3777-3788. 

The final step in the denitrification process is carried out by the soluble enzyme nitrous oxide reductase (NoS). This enzyme has been isolated from a number of sources, and is unusual in a number of ways. In most cases, it is a homodimer of ca. 74 kDa subunits with ca. 4 Cu/subunit, but the enzyme is bright purple or pink as isolated, depending on conditions, and becomes the typical blue color expected for copper proteins only after reduction with dithionite. A variety of spectroscopic studies strongly suggest that the enzyme contains at least one mixed-valent, thiolate-bridged Cu(l)—Cu(ll) unit that may well be similar to a binuclear copper center in cytochrome c oxidase (23). The reaction catalyzed by the enzyme is deceptively simple ... 

---