Siroheme

Siroheme was first found as the prosthetic group of assimilatory sulfite reductase from E. coli (Murphy et al.,1973). The heme is a complex of sirohydrochlorin with iron (Fig. 4.3), and sirohydrochlorin is biosynthesized from uroporphyrinogen III via dihydrosirohydrochlorin (Fig. 4.4). 

Recently, some bacteria have been found which oxidize organic or inorganic compounds with arsenate or selenate (arsenic and selenium respiration) (Stolz and Oremland, 1999). Although many of these bacteria are heterotrophic, Desulfovibrio auripigmentum oxidizes hydrogen with arsenate (and with sulfate, sulfite, thiosulfate, and fumarate). 

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In 1973, the first naturally occurring isobacteriochlorin, iron-containing siroheme, was isolated1 from a sulfite reductase of Escherichia coli. Later it was also discovered in sulfite and nitrite reductases of numerous bacteria and plants.2 Iron-free sirohydrochlorins (also called factor II) were discovered in vitamin B12 producing bacteria.3-4 Together with factor III. a sirohydrochlorin methylated in the 20-position, the reduced forms of factor II and factor III were identified as biosynthetic intermediates in the biosynthesis of vitamin B12.5... 

The important biological role of the isobacteriochlorins has decisively influenced the development of synthetic approaches leading to the isobacteriochlorin class of compounds. All of the naturally occurring isobacteriochlorins contain geminally dialkylated structural parts in the saturated pyrrole rings, which require special approaches for their synthesis. Until the discovery of siroheme and sirohydrochlorin, this structural element could only be found in vitamin B,2. Using the synthetic potential, which was invented during numerous syntheses of... 

Desulforubidin was found in strains of the Desulfomicrobium genus and has been described as the sulfite reductase of this genus. The subunit composition and molecular mass are similar to what was observed for desulfoviridin. However, in desulforubidin all sirohydrochlorins are metalated as proved by Mossbauer spectroscopy (152). The as-isolated protein contains four [4Fe-4S] clusters two of them are exchange-coupled to two paramagnetic sirohemes. 

Low-spin sulfite reductase were isolated from D. vulgaris (160), De-sulfuromonas acetoxidans (161), and Methanosarcina barker) (DSM 800) (162). The D. vulgaris protein has a molecular mass of 27 kDa and contains a single [4Fe-4S] cluster and one siroheme. The EPR spectrum shows a rhombic signal with g values at 2.44, 2.36, and 1.77, characteristic of a ferriheme low-spin system. This is a unique... 

Arendsen, A.F., Verhagen, M.F.J.M., Wolbert, R.B.G., Pierik, A.J., Stams, A.J.M., Jetten, M.S.M., and Hagen, W.R. 1993. Thedissimilatory sulfite reductase fromDesulfosarcina variabilis is a desulforubidin containing uncoupled metalated siroheme and S = 9/2 iron-sulfur clusters. Biochemistry 32 10323-10330. 

Pierik, A.J. and Hagen, W.R. 1991. S = 9/2 EPR signals are evidence against coupling between the siroheme and the Fe/S cluster prosthetic groups in Desulfovibrion vulgaris (Hildenborough) dissimilatory sulfite reductase. European Journal of Biochemistry 195 505-516. 

Although electron transfers in biological systems are generally expected to be non-adiabatic, it is possible for some intramolecular transfers to be close to the adiabatic limit, particularly in proteins where several redox centers are held in a very compact arrangement. This situation is found for example in cytochromes C3 of sulfate-reducing bacteria which contain four hemes in a 13 kDa molecule [10, 11], or in Escherichia coli sulfite reductase where the distance between the siroheme iron and the closest iron of a 4Fe-4S cluster is only 4.4 A [12]. It is interesting to note that a very fast intramolecular transfer rate of about 10 s was inferred from resonance Raman experiments performed in Desulfovibrio vulgaris Miyazaki cytochrome Cj [13]. 

Nitrite reductase (NAD(P)H) [EC 1.6.6.4] catalyzes the reaction of three NAD(P)H with nitrite to yield three NAD(P)+, NH4OH, and water. Cofactors for this enzyme include FAD, non-heme iron, and siroheme. (2) Nitrite reductase (cytochrome) [EC 1.7.2.1] is a copper-depen-dent system that catalyzes the reaction of nitric oxide with two ferricytochrome c and water to produce nitrite and two ferrocytochrome c. (3) Ferredoxin-nitrite reductase [EC 1.7.7.1], a heme- and iron-dependent enzyme, catalyzes the reaction of ammonia with three oxidized ferredoxin to produce nitrite and three reduced ferredoxin. (4) Nitrite reductase [EC 1.7.99.3] is a copper- and FAD-dependent enzyme that catalyzes the reaction of two nitric oxide with an acceptor substrate and two water to produce two nitrite and the reduced acceptor. 

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. 

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... 

A, while the distance from the siroheme iron to the nearest cluster iron atom is 4.4 A. One of the cluster sulfur atoms is in van der Waals contact with the siroheme ring. The X-ray structure and spectroscopic experiments (Madden et al., 1989) suggest that a cysteine thiol sulfur serves as the bridging ligand between the cluster and the siroheme. The sixth coordination site of the siroheme appears to be vacant and solvent exposed. This site presumably represents the location of substrate binding to sulfite reductase. 

The mechanism of substrate reduction by sulfite reductase has not been established. The close contact between the 4Fe 4S cluster and the siroheme could provide an efficient pathway for multielectron transfer from the enzyme to the substrate (McRee et al., 1986). Of special significance is the possibility that the cluster—siroheme overlap could stabilize high-oxidation states of the siroheme that might be involved in the catalytic mechanism. With the availability of genetic, biochemical, spectroscopic, and crystallographic approaches, it is anticipated that rapid progress will be made in working out the details of substrate reduction by sulfite reductase. 

Siroheme-comaining enzymes, 47 92 Site-directed mutagenesis dioxygenase, 47 112-113 EPR, 47 450-461... 

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. 

Both siroheme enzymes form ferroheme-NO complexes in which the g value anisotropy appears somewhat smaller than in the corresponding complexes of most other enzymes. The EPR spectra of the complexes somewhat resemble the spectra of the high-temperature myoglobin-NO complexes. The hyperfine splitting from the NO nitrogen nucleus is evident at intermediate g values but is not well resolved. These enzymes are capable of reducing NO to ammonia if supplied with low potential reducing equivalents. Other heme proteins also catalyze oxidation reduction reactions with NO. 

Krueger, R. J., and Siegel, L. M. (1982). Spinach siroheme enzymes Isolation and characterization of ferredoxin-sulfite reductase and comparison of properties with ferre-doxin-nitrite reductase. Biochemistry 21, 2892-2904. 

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. 

Vega, J. M., Garrett, R. H., and Siegel, L. M. (1975). Siroheme A prosthetic group of the Neurospora crassa assimilatory nitrite reductases. ]. Biol. Chem. 250, 7980-7989. 

Some metalloflavoproteins contain heme groups. The previously mentioned flavocytochrome b2 of yeast is a 230-kDa tetramer, one domain of which carries riboflavin phosphate and another heme. A flavocytochrome from the photosynthetic sulfur bacterium Chromatium (cytochrome c-552)279 is a complex of a 21-kDa cytochrome c and a 46-kDa flavoprotein containing 8a-(S-cysteinyl)-FAD. The 670-kDa sulfite reductase of E. coli has an a8P4 subunit structure. The eight a chains bind four molecules of FAD and four of riboflavin phosphate, while the P chains bind three or four molecules of siroheme (Fig. 16-6) and also contain Fe4S4 clusters.280 281 Many nitrate and some nitrite reductases are flavoproteins which also contain Mo or... 

Figure 16-6 Structures of isobacteriochlorin prosthetic groups. (A) Siroheme from nitrite and sulfite reductases (B) acrylochlorin heme from dissimilatory nitrite reductases of Pseudomonas and Paracoccus.

Heme d is a chlorin,85 as is acrylochlorin heme from certain bacterial nitrite reductases (Fig. 16-6).86 87 Siroheme (Fig. 16-6), which is found in both nitrite and sulfite reductases of bacteria (Chapter 24) 38/89 is an isobacteriochlorin in which both the A and B rings are reduced. It apparently occurs as an amide siroamide (Fig. 16-6) in Desulfovibrio.90 Heme of nitrite reductases of denitrifying bacteria is a dioxo-bacteriochlorin derivative (Fig. 16-6).91 92... 

Iron-sulfur clusters are found in flavoproteins such as NADH dehydrogenase (Chapter 18) and trimethylamine dehydrogenase (Fig. 15-9) and in the siroheme-containing sulfite reductases and nitrite reductases.312 These two reductases are found both in bacteria and in green plants. 

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... 

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