Abstract
Three cytochromes of the thiosulfate-utilizing green sulfur bacterium Chlorobium vibrioforme f. thiosulfatophilum were highly purified by ion exchange column chromatography and ammonium sulfate fractionation. All three cytochromes are located in the soluble fraction. Cytochrome c-551 (highest purity index obtained: A280/A416=0.39) shows maxima at 551 nm (α-band), 521 nm (β-band), and 416 nm (γ-band) for the reduced form. This cytochrome is an acidic protein with a molecular weight of 32,000, a redox potential of 150 mV, and an isoelectric point at pH 6.0. Cytochrome c-553 (highest purity index obtained: A280/A417=0.8) is also an acidic protein with maxima at 553,5 nm, 523,5 nm and 417 nm for the reduced form, a molecular weight of 63,000, a redox potential of 90 mV, an isoelectric point at pH 6.3, and it contains FAD as flavin component. It is autoxidizable and participates in sulfide oxidation, but cannot catalyze the reverse reaction. The cytochrome c-555 (highest purity index obtained: A280/A418=0.16) is a small basic protein with maxima at 555 nm, 523 nm and 418 nm (reduced form), a molecular weight of 12,500, an isoelectric point between pH 10 and 10.5, and a redox potential of 155 mV. The ratio of the cytochrome contents to each other is constant and does not change when the organism has only thiosulfate or sulfide as the main electron donor in the medium.
The soluble fraction further contains the non-heme ironcontaining proteins rubredoxin and ferredoxin. The anaerobic sulfide oxidation in a growing culture of Chlorobium vibrioforme f. thiosulfatophilum is accompanied by a rapid formation of thiosulfate, which is only utilized when sulfide is no longer available, while the elemental sulfur concentration increases constantly until thiosulfate is consumed.
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Abbreviations
- C:
-
Chlorobium
- SDS:
-
sodium dodecylsulfate
- HIPIP:
-
high-potential-iron-sulfur-protein
References
Appaji N, Felton SP, Huennekens FM (1967) Quantitative determination of mitochondrial flavins. In: RW Estabrock, ME Pullmann (eds) Methods in enzymology, vol 10. Academic Press, New York, pp 494–499
Appelt N, Weber H, Wieluch S, Knobloch K (1979) Über das System Thiosulfat-Cytochrom c-Oxidoreduktase aus Rhodopseudomonas palustris. Ber Dt Bot Ges 92:365–378
Bartlett JK, Skoog DA (1954) Colorimetric determination of elemental sulfur in hydrocarbons. Anal Chem 26:1003–1011
Bartsch RG, Meyer TE, Robinson AB (1968) Complex c-type cytochromes with bound flavin. In: K Okunuki, MD Kamen, I Sekuzu (eds) Structure and function of cytochromes. University of Tokyo Press, Tokyo, pp 443–451
Bartsch RG (1971) Cytochromes: Bacterial. In: SP Collowick, NO Kaplan (eds) Methods in enzymology, vol 23. Academic Press, New York, pp 344–363
Bartsch RG (1978) Cytochromes. In: RK Clayton, WR Sistrom (eds) The photosynthetic bacteria. Plenum Press, New York London, pp 249–279
Fauque G, Herve D, Le Gall J (1979) Structure function relationship in hemoproteins: the role of cytochrome c3 in the reduction of colloidal sulfur by sulfate reducing bacteria. Arch Microbiol 121:261–264
Fischer U (1977) Die Rolle von Cytochromen im Schwefelstoffwechsel phototropher Schwefelbakterien. Doctoral thesis, Univ Bonn
Fischer U, Trüper HG (1977) Cytochrome c-550 of Thiocapsa roseopersicina: Properties and reduction by sulfide. FEMS Lett 1:87–90
Fischer U, Trüper HG (1979) Some properties of cytochrome c′ and other hemoproteins of Thiocapsa roseopersicina. Curr Microbiol 3:41–44
Fukumori Y, Yamanaka T (1979a) Flavocytochrome c of Chromatium vinosum. Some enzymatic properties and subunit structure. J Biochem 85:1405–1414
Fukumori Y, Yamanaka T (1979b) A high potential nonheme iron protein (HIPIP)-linked, thiosulfate-oxidizing enzyme derived from Chromatium vinosum. Curr Microbiol 3:117–120
Gibson J (1961) Cytochrome pigments from the green photosynthetic bacterium Chlorobium thiosulfatophilum. Biochem J 79:151–158
Hatchikian EC, Le Gall J, Bruschi M, Dubourdieu M (1972) Regulation of the reduction of sulfite and thiosulfate by ferredoxin, flavodoxin and cytochrome cc′3 in extracts of the sulfate reducer Desulfovibrio gigas. Biochim Biophys Acta 258:701–708
Kilgour GL, Felton SP, Huennekens FM (1957) Paper chromatography of flavins and flavin nucleotides. J Am Chem Soc 79:2254–2256
Knobloch K, Schmitt W, Schleifer G, Appelt N, Müller H (1981) On the enzymatic system thiosulfate-cytochrome c-oxidoreductase. In: H Bothe, A Trebst (eds) Biology of inorganic nitrogen and sulfur. Springer, Berlin Heidelberg New York, pp 359–365
Kusai A, Yamanaka T (1973a) A novel function of cytochrome c (555, Chlorobium thiosulfatophilum) in oxidation of thiosulfate. Biochem Biophys Res Comm 51:107–112
Kusai A, Yamanaka T (1973b) The oxidation mechanism of thiosulfate and sulfide in Chlorobium thiosulfatophilum. Roles of cytochrome c-551 and cytochrome c-553. Biochim Biophys Acta 325:304–314
Kusai A, Yamanaka T (1973c) Cytochrome c (553, Chlorobium thiosulfatophilum) is a sulfide-cytochrome c reductase. FEBS Lett 34:235–237
Lowry HD, Rosebrough NJ, Farr AL, Randall RJ (1951) Protein measurement with the Folin reagent. J Biol Chem 193:265–275
Lyons D, Nickless G (1968) The lower oxy-acids of sulphur. In: G Nickles (ed) Inorganic sulphur chemistry. Elsevier Publishing Company, Amsterdam, pp. 509–533
Meyer TE, Bartsch RG, Cusanovich MA, Mathewson JH (1968) The cytochromes of Chlorobium thiosulfatophilum. Biochim Biophys Acta 153:854–861
Meyer TE, Sharp JJ, Bartsch RG (1971) Isolation and properties of rubredoxin from the photosynthetic green sulphur bacteria. Biochim Biophys Acta 234:266–269
Moriarty DJW, Nicholas DJD (1970) Electron transfer during sulphide and sulphite oxidation by Thiobacillus concretivorus. Biochim Biophys Acta 216:130–138
Pachmayr F (1960) Vorkommen und Bestimmung von Schwefelverbindungen in Mineralwasser. Doctoral thesis, Univ Munich
Paschinger H, Paschinger J, Gaffron H (1974) Photochemical disproportionation of sulfur by Chlorobium limicola f. thiosulfatophilum. Arch Microbiol 96:341–351
Pfennig N (1965) Anreicherungskulturen für rote und grüne Schwefelbakterien. In: HG Schlegel, E Kröger (es) Anreicherungskulturen und Mutantenauslese. Fischer, Stuttgart, pp 179–189, 503–504
Pfennig N, Lippert KD (1966) Über das Vitamin B12-Bedürfnis phototropher Schwefelbakterien. Arch Mikrobiol 55:245–256
Roy AB, Trudinger PA (1970) The biochemistry of inorganic compounds of sulfur. Cambridge Univ Press, pp 43–44
Schedel M (1978) Untersuchungen zur anaeroben Oxidation reduzierter Schwefel verbindungen durch Thiobacillus denitrificans, Chromatium vinosum und Chlorobium limicola. Doctoral thesis, Univ Bonn
Schedel M, Trüper HG (1979) Purification of Thiobacillus denitrificans siroheme sulfite reductase and investigation of some molecular and catalytic properties. Biochim Biophys Acta 568:454–467
Steinmetz MA, Fischer U (1981) Cytochromes of the non-thiosulfateutilizing green sulfur bacterium Chlorobium limicola. Arch Microbiol 130:31–37
Trudinger PA (1961a) Thiosulfate oxidation and cytochromes in Thiobacillus X. I. Fractionation of bacterial extracts and properties of cytochromes. Biochem J 78:673–680
Trudinger PA (1961b) Thiosulfate oxidation and cytochromes in Thiobacillus X. II. Thiosulfate-oxidizing enzymes. Biochem J 78:681–686
Trüper HG, Schlegel HG (1964) Sulphur metabolism in Thiorhodaceae. I. Quantitative measurements on growing cells of Chromatium okenii. Antonie van Leeuwenhoek J Microbiol Serol 30:225–238
Trüper HG, Rogers LH (1971) Purification and properties of adenylylsulfate reductase from the phototrophic sulfur bacterium Thiocapsa roseopersicina. J Bacteriol 108:112–1121
Urban PJ (1961) Colorimetry of sulfur anions. Part I. An improved colorimetric method for determination of thiosulfate. Z Analyt Chem 179:415–422
Weber K, Osborne M (1969) The reliability of molecular weight determinations by dodecylsulfate-polyacrylamide-gel electrophoresis. J Biol Chem 244:4406–4412
Yamanaka T, Kusai A (1976) The function and some molecular features of cytochrome c-553 derived from Chlorobium thiosulfatophilum. In: TP Singer (ed) Flavins and flavoproteins. Elsevier Scientific Publishing Company, Amsterdam, pp 292–301
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Steinmetz, M.A., Fischer, U. Cytochromes of the green sulfur bacterium Chlorobium vibrioforme f. thiosulfatophilum. Purification, characterization and sulfur metabolism. Arch. Microbiol. 131, 19–26 (1982). https://doi.org/10.1007/BF00451493
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DOI: https://doi.org/10.1007/BF00451493