Reaction of sulfenyl halides with cytochrome <i>c</i>. A novel method for heme cleavage

Fontana, Angelo; Veronese, Francesco M.; Boccù, E

doi:10.1016/0014-5793(73)80756-2

Cited by 44 publications

(22 citation statements)

References 15 publications

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“…Covalently bound c-type heme was removed using 2-nitrophenylsulfenyl chloride (Sigma-Aldrich, St. Louis, MO) according to the method of Fontana et al (7). SDS-PAGE with a SHreducing reagent was used to determine the molecular mass of the protein whose heme had been removed.…”

Section: Methodsmentioning

confidence: 99%

Isolation of a Multiheme Protein with Features of a Hydrazine-Oxidizing Enzyme from an Anaerobic Ammonium-Oxidizing Enrichment Culture

Shimamura

Nishiyama

Shigetomo

et al. 2007

Appl Environ Microbiol

154

109

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A multiheme protein having hydrazine-oxidizing activity was purified from enriched culture from a reactor in which an anammox bacterium, strain KSU-1, was dominant. The enzyme has oxidizing activity toward hydrazine but not hydroxylamine and is a 130-kDa homodimer composed of a 62-kDa polypeptide containing eight hemes. It was therefore named hydrazine-oxidizing enzyme (HZO). With cytochrome c as an electron acceptor, the V max and K m for hydrazine are 6.2 ؎ 0.3 mol/min · mg and 5.5 ؎ 0.6 M, respectively. Hydrazine (25 M) induced an increase in the proportion of reduced form in the spectrum, whereas hydroxylamine (500 M) did not. Two genes coding for HZO, hzoA and hzoB, were identified within the metagenomic DNA from the culture. The genes encode the same amino acid sequence except for two residues. The sequences deduced from these genes showed low-level identities (<30%) to those of all of the hydroxylamine oxidoreductases reported but are highly homologous to two hao genes found by sequencing the genome of "Candidatus Kuenenia stuttgartiensis" (88% and 89% identities). The purified enzyme might therefore be a novel hydrazine-oxidizing enzyme having a critical role in anaerobic ammonium oxidation.The anaerobic ammonium oxidation (anammox) process is a fascinating subject of study, not only because it is an innovative technological advance for the removal of nitrogenous contaminants from wastewater (14,17,27), but also because it includes interesting and novel biochemical reactions. In the anammox reaction, ammonium and nitrite contribute in equimolar amounts to the formation of dinitrogen gas (N 2 ) according to the following equation (26): NH 4 ϩ ϩ NO 2 Ϫ 3 N 2 ϩ 2H 2 O; G°ϭ Ϫ358 kJ/mol ammonium. This biological reaction is performed under anoxic conditions by novel autotrophic bacteria, which have not yet been isolated but which are identified as deeply branching planctomycetes, based on nucleotide sequences of 16S rRNA genes (17,22,26). The first anammox bacterium to be identified, discovered in The Netherlands, has been provisionally named "Candidatus Brocadia anammoxidans" (15). An increasing number of 16S rRNA gene sequences from other putative anaerobic ammonium-oxidizing bacteria have been reported to date from laboratories (6,21,29,31) and from natural marine habitats such as the Black Sea (4, 5, 13, 16).We designed a continuous-flow reactor shielded from light in which to enrich cultures for bacteria having the ability to catalyze the anammox reaction from a seed of denitrifying sludge originating in Kumamoto City, Japan. In the reactor, the culture turned a distinctly red color and yielded large amounts of sludge that could remove ammonium and nitrite with a relatively low production of nitrate at ratios strongly resembling that of the anammox reaction (20). To investigate the bacterial composition of the enriched culture community, 16S rRNA gene sequences were amplified by PCR and cloned. The dominant clones had an identical sequence, which has 92.2% identity with the 16S rRNA gene of "Candidatus B...

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Section: Methodsmentioning

confidence: 99%

Isolation of a Multiheme Protein with Features of a Hydrazine-Oxidizing Enzyme from an Anaerobic Ammonium-Oxidizing Enrichment Culture

Shimamura

Nishiyama

Shigetomo

et al. 2007

Appl Environ Microbiol

154

109

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“…As reported by Erickson and Hooper (1), the prosthetic group was not extracted with acidic acetone. We tried further to remove the prosthetic group by treating the cytochrome with Ag 2 SO 4 (15), o-nitrophenylsulfenyl chloride (16), SDS-urea-mercaptoethanol, and pronase. However, the prosthetic group was not extracted with acetone or ethyl acetate at acidic pH from the cytochrome thus treated.…”

Section: Molecularmentioning

confidence: 99%

Cytochrome P-460 of Nitrosomonas europaea: Further Purification and Further Characterization

Numata

Saito

Yamazaki

et al. 1990

The Journal of Biochemistry

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Cytochrome P-460 of Nitrosomonas europaea [Erickson, R.H. and Hooper, A.B. (1972) Biochim. Biophys. Acta 275, 231-244] was further purified to an electrophoretically homogeneous state. The cytochrome molecule was composed of three molecules of subunits with Mr of 17,300-18,500, and contained three atoms of iron, which seemed to be heme iron, and six cysteine residues, but did not contain nonheme iron or inorganic sulfide. The cytochrome showed absorption peaks at 460 and 688 nm with a broad shoulder at 635 nm in the reduced form. The ESR spectrum of ferricytochrome P-460 showed signals at g = 5.91, 5.63, and 1.99, indicating that the protein was a high spin hemoprotein. The heme of the cytochrome was not cleaved by the methods which were available for cleavage of heme c. The pyridine ferrohemochrome of the hemoprotein did not show the distinct alpha and beta peaks which are shown by the ferrohemochromes of many other cytochromes so far known. The N-terminal amino acid sequence of cytochrome P-460 differed from that of hydroxylamine oxidoreductase. Therefore, cytochrome P-460 did not seem to be the solubilized P-460 moiety of hydroxylamine oxidoreductase, in agreement with the finding by D.J. Miller et al. [J. Gen. Microbiol. 130, 3049-3054 (1984)]. However, cytochrome P-460 had several enzymatic activities which hydroxylamine oxidoreductase showed. Although most of the activities of the cytochrome were lower than the corresponding activities of the oxidoreductase, the hydroxylamine-cytochrome c-552 reductase activity of the cytochrome was about 5-times as high as that of the oxidoreductase.

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“…In order to release the covalently bound heme, the isolated complex I11 (120 mg) was treated with nitrophenylsulphenyl chloride in 70% formic acid instead of acetic acid [45]. The mixture was incubated for 20 min at 37°C with a gentle stirring, then diluted threefold with water and extracted four times with ethyl acetate using low-speed centrifugation.…”

Section: Preparation Of Cm-cytochrome C1mentioning

confidence: 99%

An atypical heme‐binding structure of cytochrome c₁ of Euglena gracilis mitochondrial complex III

Mukai¹,

Yoshida²,

Toyosaki³

et al. 1989

European Journal of Biochemistry

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Complex III was purified from submitochondrial particles prepared from Euglena gracilis. The purified complex consisted of 10 subunits and lost antimycin sensitivity. The Euglena complex III showed an atypical difference absorption spectrum for cytochrome c1 with its α‐band maximum at 561 nm. The pyridine ferrohemochrome prepared from covalently bound heme in the Euglena complex III had an α‐peak at 553 nm. This wavelength is the same as that of pyridine ferrohemochrome prepared from Euglena mitochondrial cytochrome c (c‐558), the heme of which is linked to only a single cysteine residue through a thioether bond. Cytochrome c1 which was a heme‐stained subunit with a molecular mass of 32.5 kDa was isolated from the purified complex III and its N‐terminal sequence of 46 amino acids was determined. On the basis of apparent homologies to cytochromes c1 from other sources, this sequence included the heme‐binding region. However, the amino acid at position 36, corresponding to the first cysteine involved in heme linkage in other cytochromes c1, was phenylalanine. Position 39, corresponding to the second cysteine, was not identified despite the treatment for removal of the heme and carboxymethylation of the expected cysteine. The unidentified amino acid is assumed to be a derivative of cysteine to which the heme is linked through a single thioether bond. The histidine‐40 corresponding to the probable fifth ligand for heme iron was conserved in Euglena cytochrome c1.

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Reaction of sulfenyl halides with cytochrome c. A novel method for heme cleavage

Cited by 44 publications

References 15 publications

Isolation of a Multiheme Protein with Features of a Hydrazine-Oxidizing Enzyme from an Anaerobic Ammonium-Oxidizing Enrichment Culture

Isolation of a Multiheme Protein with Features of a Hydrazine-Oxidizing Enzyme from an Anaerobic Ammonium-Oxidizing Enrichment Culture

Cytochrome P-460 of Nitrosomonas europaea: Further Purification and Further Characterization

An atypical heme‐binding structure of cytochrome c₁ of Euglena gracilis mitochondrial complex III

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Reaction of sulfenyl halides with cytochrome c. A novel method for heme cleavage

Cited by 44 publications

References 15 publications

Isolation of a Multiheme Protein with Features of a Hydrazine-Oxidizing Enzyme from an Anaerobic Ammonium-Oxidizing Enrichment Culture

Isolation of a Multiheme Protein with Features of a Hydrazine-Oxidizing Enzyme from an Anaerobic Ammonium-Oxidizing Enrichment Culture

Cytochrome P-460 of Nitrosomonas europaea: Further Purification and Further Characterization

An atypical heme‐binding structure of cytochrome c1 of Euglena gracilis mitochondrial complex III

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An atypical heme‐binding structure of cytochrome c₁ of Euglena gracilis mitochondrial complex III