Summary
A cytochrome bc 1-complex of Rs. rubrum was isolated and the three subunits were purified to homogeneity. The N-terminal amino acid sequence of the purified subunits was determined by automatic Edman degradation. The pet genes of Rhodospirillum rubrum coding for the three subunits of the cytochrome bc 1-complex were isolated from a genomic library of Rs. rubrum using oligonucleotides specific for conserved regions of the subunits from other organisms and a heterologous probe derived from the genes for the complex of Rb. capsulatus. The complete nucleotide sequence of a 5500 by SalI/SphI fragment is described which includes the pet genes and three additional unidentified open reading frames. The N-terminal amino acid sequence of the isolated subunits was used for the identification of the three genes. The genes encoding the subunits are organized as follows: Rieske protein, cytochrome b, cytochrome c 1. Comparison of the N-terminal protein sequences with the protein sequences deduced from the nucleotide sequence showed that only cytochrome c 1 is processed during transport and assembly of the three subunits of the complex. Only the N-terminal methionine of the Rieske protein is cleaved off. The similarity of the deduced amino acid sequence of the three subunits to the corresponding subunits of other organisms is described and implications for structural features of the subunits are discussed.
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Abbreviations
- BSA:
-
bovine serum albumin
- SDS:
-
sodium dodecylsulphate
- Rs:
-
Rhodospirillum
- Rb:
-
Rhodobacter
- Pc:
-
Paracoccus
- Rps:
-
Rhodopseudomonas
References
Argos P, Mohana Rao JK, Hargrave PA (1982) Structural prediction of membrane-bound proteins. Eur J Biochem 128:565–575
Brasseur R (1988) Calculation of the three-dimensional structure of Saccharomyces cerevisiae cytochrome b inserted in a lipid matrix. J Biol Chem 263:12571–12575
Collins MLP, Niederman RA (1976) Membranes of Rhodospirillum rubrum: Physicochemical properties of chromatophore fractions isolated from osmotically and mechanically disrupted cells. J Bacteriol 126:1326–1338
Cramer WA, Black MT, Widger WR, Girvan ME (1987) In: Barber J (ed) The light reactions. Elsevier, Amsterdam, pp 447–493
Crofts A, Robinson H, Andrews K, Van Doren S, Berry E (1987) Catalytic sites for reduction and oxidation of quinones. In: Papa S, Chance B, Ernster L (eds) Cytochrome systems. Plenum Press, New York, pp 617–624
Daldal F, Tokito MK, Davidson E, Faham M (1989) Mutations conferring resistance to quinol oxidation (Qz)-inhibitors of the cytochrome bc1 complex of Rhodobacter capsulatus. EMBO J 8:3951–3961
Davidson E, Daldal F (1987) Primary structure of the bc1 complex of Rhodopseudomonas capsulata. Nucleotide sequence of the pet operon encoding the Rieske cytochrome b, and cytochrome c1 apoproteins. J Mol Biol 195:13–24
Davis DJ, Frame MK, Johnson DA (1988) Resonance Raman spectroscopy indicates a lysine as the sixth iron ligand in cytochrome f. Biochem Biophys Acta 936:61–66
Di Rago JP, Colson AM (1988) Molecular basis for resistance to antimycin and diuron, Q-cycle inhibitors acting at the Qi site in the mitochondrial ubiquinol-cytochrome c reductase in Saccharomyces cerevisiae. J Biol Chem 263:12564–12570
Di Rago JP, Coppee JY, Colson AM (1989) Molecular basis for resistance to myxothiazol, mucidin (strobilurin a), and stigmatellin. J Biol Chem 264:14543–14548
Eisenberg D, Schwarz E, Komarony M, Wall R (1984) Analysis of membrane and surface protein sequences with the hydrophobic moment plot. J Mol Biol 179:125–142
Falk G, Hampe A, Walker E (1985) Nucleotide sequence of the Rhodospirillum rubrum atp operon. Biochem J 228:391–407
Gabellini N (1988) Organization and structure of the genes for the cytochrome b/c1 complex in purple photosynthetic bacteria. A phylogenetic study describing the homology of the b/c1, subunits between prokaryotes, mitochondria and chloroplasts. J Bioenerg Biomembr 20:59–83
Gabellini N, Harnisch U, McCarthy JEG, Hauska G, Sebald W (1985) Cloning and expression of the fbc operon encoding the FeS protein, cytochrome b and cytochrome c1 from the Rhodopseudomonas sphaeroides b/c1 complex. EMBO J 4:549–553
Gabellini N, Sebald W (1986) Nucleotide sequence and transcription of the fbc operon from Rhodopseudomonas sphaeroides. Eur J Biochem 154:569–579
Garger SJ, Griffith OM, Grill LK (1983) Rapid purification of plasmid DNA by single centrifugation in a two-step cesium chloride-ethidium bromide gradient. Biochem Biophys Res Commun 117:835–842
Gibson J, Stackebrant E, Zablen LB, Gupta R, Woese CR (1979) A phylogenetic analysis of the purple photosynthetic bacteria. Curr Microbiol 3:59–64
Gierasch LM (1989) Signal sequences. Biochemistry 28:923–930
Grossberger D (1987) Minipreps of DNA from bacteriophage lambda. Nucleic Acids Res 15:6737
Harnisch U, Weiss H, Sebald W (1985) The primary structure of the iron-sulfur subunit of ubiquinol-cytochrome c reductase from Neurospora, determined by cDNA and gene sequencing. Eur J Biochem 149:95–99
Hartl FU, Schmidt B, Wachter E, Weiss H, Neupert W (1986) Transport into mitochondria and intramitochondrial sorting of the Fe/S protein of ubiquinol-cytochrome c reductase. Cell 47:939–951
Hauska G, HurtE, Gabellini G, Lockau W (1983) Comparative aspects of quinol-cytochrome c/plastocyanin oxidoreductases. Biochem Biophys Acta 726:97–133
Hauska G, Nitschke W, Herrmann RG (1988) Amino acid identities in the three redox center carrying polypeptides of cytochrome bc1/b6f-complexes. J Bioenerg Biomembr 20:211–228
Henikoff S (1987) Unidirectional digestion with Exonuclease III in DNA sequence analysis. Methods Enzymol 155:156–165
Howell N, Gilbert K (1988) Mutational analysis of the mouse mitochondrial cytochrome b gene. Mol Biol 203:607–618
Kriauciunas A, Yu L, Yu CA, Wynn RM, Knaff DB (1989) The Rhodospirillum rubrum cytochrome bc1 complex: peptide composition, prosthetic group content and quinone binding. Biochem Biophys Acta 976:70–76
Kurowski B, Ludwig B (1987) The genes of the Paracoccus denitrificans bc1 complex. J Biol Chem 262:13805–13811
Leustek T, Hartwig R, Weissbach H, Brot N (1988) Regulation of ribulose biphosphate carboxylase expression in Rhodospirillum rubrum: characteristics of mRNA synthesized in vivo and in vitro. J Bacteriol 170:4065–4071
Ljungdahl PO, Pennoyer JD, Robertson DE, Trumpower BL (1987) Purification of highly active cytochrome bc1 complexes from phylogenetically diverse species by a single chromatographic procedure. Biochem Biophys Acta 891:227–241
Majewski C (1989) The cytochrome bc1 complex of Rhodospirillum rubrum. Isolation of the complex and the coding genes. Thesis, Bochum
Malkin R (1988) Structure function studies of photosynthetic cytochrome bc1 and cytochrome b6 complexes. ISI Atlas of Science. Biochemistry: 57–64
Maniatis T, Fritsch EF, Sambrook J (1982) Molecular cloning: A laboratory manual. Cold Spring Harbor Laboratory, Cold Spring Harbor, New York
Omerod JG, Omerod KS, Gest H (1961) Light-dependent utilization of organic compounds and photoproduction of molecular hydrogen 18 by photosynthetic bacteria: relationships with nitrogen metabolism. Arch Biochem Biophys 94:449–463
Randall LL, Hardy SJS (1989) Unity in function in the absence of consensus in sequence: role of leader peptides in export. Science 243:1156–1159
Sanger F, Nicklen S, Coulson AR (1977) DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci USA 74:5463–5467
Saraste M (1984) Localisation of haem-binding sites in the mitochondrial cytochrome b. FEBS Lett 166:367–372
Schägger H, Borchart U, Machleidt W, Link TA, Von Jagow G (1987) Isolation and amino acid sequence of the Rieske ironsulfur protein of beef heart ubiquinol cytochrome c reductase. FEBS Lett 219:161–168
Silhavy TJ, Berman ML, Enquist LW (1984) Experiments with gene fusions. Cold Spring Harbor Laboratory, Cold Spring Harbor, New York
Tervoort MJ, Van Gelder BF (1983) The interaction between heme and protein in cytochrome c1. Biochem Biophys Acta 722:137–143
Thöny-Meyer L, Stax D, Hennecke H (1989) An unusual gene cluster for the cytochrome bc1 complex in Bradyrhizobium japonicum and its requirement for effective root nodule symbiosis. Cell 57:683–697
Van der Wal HN, Van Grondelle R (1983) Flash induced electron transport in b- and c-type cytochromes in Rhodospirillum rubrum. Evidence for a Q-cycle. Biochem Biophys Acta 725:94–103
Verbist J, Lang F, Gabellini N, Oesterhelt D (1989) Cloning and sequencing of the fbcF, B and genes encoding the cytochrome b/c1 complex from Rhodopseudomonas viridis. Mol Gen Genet 219:445–452
Weiss H, Leonard K, Neupert W (1990) Puzzling subunits of mitochondrial cytochrome reductase. Trends Biochem Sci 15:178–180
Widger WR, Cramer WA, Hermann RG, Trebst A (1984) Sequence homology and structural similarity between cytochrome b of mitochondrial complex III and the chloroplast b6-f complex: Position of the cytochrome b hemes in the membrane. Proc Natl Acta Sci USA 81:674–678
Wynn RM, Gaul DF, Choi WK, Shaw RW, Knaff DB (1986) Isolation of cytochrome bc1 complexes from the photosynthetic bacteria Rhodopseudomonas viridis and Rhodospirillum rubrum. Photosyn Res 9:181–195
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Communicated by J. Lengeler
The nucleotide sequence reported in this paper has been submitted to the GenBank™/EMBL Data Bank with accession number X55387
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Majewski, C., Trehst, A. The pet genes of Rhodospirillum rubrum: cloning and sequencing of the genes for the cytochrome bc 1-complex. Molec. Gen. Genet. 224, 373–382 (1990). https://doi.org/10.1007/BF00262431
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DOI: https://doi.org/10.1007/BF00262431