Siroheme-containing enzymes

Vega, J. M., and Kamin, H. (1977). Spinach nitrite reductase. Purification and properties of a siroheme-containing iron-sulfur enzyme. J. Biol. Chem. 252, 896-909. 

There are four different types of nitrite reductases the copper-containing protein Copper Enzymes in Denitrification and cytochrome cd perform a one-electron rednetion of nitrite to nitric oxide, and are involved in denitrification " the siroheme-containing protein and the cytochrome c ititrite reductase (cNiR) both perform the complete, six-electron reduction, of nitrite to ammonia. The cNiR is present in the y, 5 or e-subclasses of proteobacteria, and is encoded by the nrf operon (nitrite reduction with /ormate), which has different gene composition in the different classes of bacteria, having in common only the gene for the catalytic subunit, ntfA. 

There are two classes of nitrite reductases those involved in denitrification, which reduce nitrite to gaseous nitrogen oxides and the assimilatory and dis-similatory enzymes, which reduce nitrite directly to ammonia [125, 126]. Two types of denitrifying enzymes have been described, those containing hemes c and dx and those which contain only copper. There are also two kinds of as-similatory/dissimilatory enzymes the siroheme containing nitrite reductases like that of E. coli, which is used for detoxification of nitrite from the cytoplasm and is not coupled to energy conservation and the heme c nitrite reductases, which are usually coupled to energy conservation [127]. This last type of nitrite reductase is that usually associated with strict anaerobes, so only this one will be discussed in more detail. 

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. 

Nitrite reductase and sulfite reductase are enzymes found in choroplasts and in prokaryotes that reduce nitrite to ammonia and sulfite to sulfide (Scott et al., 1978). Sulfite reductase also catalyzes reduction of nitrite at a lower rate. Both enzymes contain a siroheme prosthetic group linked to an iron-sulfur cluster. In siroheme, the porphyrinoid moiety is present in the more reduced chlorin form. Because NO lies between nitrite and ammonia in oxidation state, it is a potential intermediate. 

Spinach nitrite reductase,313 which is considered further in Chapter 24, utilizes reduced ferredoxin to carry out a six-electron reduction of N02 to NH3 or of SO-2 to S2. The 61-kDa monomeric enzyme contains one siroheme and one Fe4S4 cluster. A sulfite reductase from E. coli utilizes NADPH as the reductant. It is a large (38a4 oligomer.312 The 66-kDa a chains contain bound flavin... 

The enzymes from green plants and fungi are large multifunctional proteins,80 which may resemble assimilatory sulfite reductases (Fig. 16-19). These contain siroheme (Fig. 16-6), which accepts electrons from either reduced ferredoxin (in photosynthetic organisms) or from NADH or NADPH. FAD acts as an intermediate carrier. It seems likely that the nitrite N binds to Fe of the siroheme and remains there during the entire six-electron reduction to NH3. Nitroxyl (NOH) and hydroxylamine (NH2OH) may be bound intermediates as is suggested in steps a-c of Eq. 24-14. 

In various species of bacteria several different types of non-assimilatory nitrite reductases are found. Escherichia coli has a cytoplasmic NAD(P)H-dependent enzyme whose role seems to be detoxification of nitrite. This type of enzyme, coded for by the nirB gene, also contains siroheme as the redox active catalytic center (Cole, 1988). Additionally in E. coli, and expressed under different conditions to the cytoplasmic enzyme, is a periplasmic nitrite reductase that catalyses formation of ammonia from nitrite (Cole, 1988). This enzyme has five c-type (Figure 1) hemes per polypeptide chain one of these hemes, the catalytic site, has the unique CXXCK sequence as its attachment site (Einsle et al., 1999). Electrons reach this type of nitrite reductase, which is fairly widely distributed amongst the microbial world, from the cytoplasmic membrane electron transfer chain. The exact electron donor partner from such chains for this type of nitrite reductase is unknown (Berks et al., 1995). 

The initial six-electron oxidation of sulfide to sulfite is catalysed by a soluble, dis-similatory sulfite reductase that contains siroheme and at least one iron-sulfur center as prosthetic groups [83-85]. While similar enzymes in plants and in non-pho-totrophic bacteria usually function to reduce sulfite to sulfide in assimilatory pathways, the enzyme in photolithoautotrophically grown C. vinosum appears to function in the reverse direction, with the electrons from sulfide oxidation being delivered to an as yet unidentified acceptor. Evidence is also available for the... 

Sulfite reductases contain siroheme and iron-sulfur centers. Siroheme, also present in some nitrite reductases, is an iron tetrahydroporphyrin of the isobacteriochlorin type with eight carboxylic acid side-chains (Fig. 1). Siroheme isolated from Desulfovibrio species was found to be a monoamide, heptamethyl ester derivative, rather than the usual octamethyl ester derivative, which suggests that in these organisms an amidated form of the siroheme may be the physiologically active prosthetic group [93]. Sulfite reductases are divided into two classes, the assimilatory and the dissimilatory enzymes. The assimilatory sulfite reductases produce sulfide for use in the cell biosynthetic pathways. The dissimilatory enzymes are present in the sulfate-reducing organisms, and reduce sulfite as a respiratory substrate in a process coupled to ATP formation. 

Desulfoviridin was reported to contain a siroheme which was more than 70% demetallated [112-114], but it was described that a more careful purification of the D. vulgaris desulfoviridin yields a fully metallated enzyme [115]. However, a recent report where this purification procedure was repeated contradicts this observation [116]. 

The last three steps in the reduction of nitrate to ammonia are carried out by an enzyme called nitrite reductase. It contains one Fe2S2 center and one molecule of siroheme, a partially reduced iron porphyrin. The electron donor for each step is ferredoxin. 

The purified enzyme has a molecular weight of about 6.2 x Kf (Hucklesby et al., 1976 Ida, 1977 Vega and Kamin, 1977) and is not dissociated by SDS (Ida, 1977 Vega and Kamin, 1977) and so apparently does not contain subunits. Iron at a concentraticxi of 4.2-1.82 atoms/mol has been detected in the purified enzyme (Hucklesby et al., 1976 Ida, 1977 Vega and Kamin, 1977). These data are interpreted as indicating three atoms of iron/ mol of enzyme one iron atom is associated with a heme compound (Vega and Kamin, 1977) identified as siroheme (Murphy et al., 1974 Hucklesby et... 

Macrocyclic tetrapyrrole compounds such as heme (iron), chlorophyll (magnesium), siroheme (iron), and E12 (cobalt) contain specific metal ions at the center of their tetrapyrrole rings [17]. Metal ion chelatases can be divided into two classes based on their structural architecture. Class 1 chelatases are heteromultimeric enzymes that require three gene products for efficient catalysis [18] of the ATP-dependent chelation reaction [19]. Enzymes in this class include chlorophyll and bacteriochlorophyll magnesium chelatases [18] and aerobic cobalt chelatase (CobNST) [20]. 

Sulfite reductase, another enzyme which is not directly involved in sulfate assimilation, has also been studied in detail in spinach (Kruger and Si el, 1982). Like nitrite reductase it contains siroheme and supports both nitrite... 

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