The pet genes of Rhodospirillum rubrum: cloning and sequencing of the genes for the cytochrome bc 1-complex

Abstract

A cytochrome bc1-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 bc1-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 bp 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 c1. Comparison of the N-terminal protein sequences with the protein sequences deduced from the nucleotide sequence showed that only cytochrome c1 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.

Molecular features, processing and import of the Rieske iron-sulfur protein from potato mitochondria

“…The latter has been characterized as the last of the redox-center containing subunits of the cytochrome bcl complex from potato mitochondria. The mature iron-sulfur protein shows more than 50~o sequence identity with the mature protein from yeast and Neurospora and even 54~o sequence identity with the iron-sulfur protein from a prokaryote, Rhodospirillum rubrum [24]. This non-sulfur purple bacterium has a three subunit bcl complex and the high similarity of its ironsulfur subunit with those from fungal and mammalian mitochondria (45-50~o sequence identity) and especially with those from plant mitochondria supports the idea that there is a close phylogenetic relationship between purple bacteria and mitochondria.…”

“…To analyse the maturation of the iron-sulfur pro- [19], Neurospora [16], yeast [1], human [24] and bovine [4,28]. Identical amino acids are boxed if present in at least four organisms.…”

Characterization of the pet operon of Rhodospirillum rubrum

“…Despite functional similarity and the presence of the Rieske iron-sulfur protein, cytochrome b, and cytochrome c I in all cytochrome bcl complexes, the complexes differ with regard to their subunit composition. In photosynthetic purple sulfur bacteria these complexes contain either three (Wynn et al 1986, Cully et al 1989, Kriauciunas et al 1989, Purvis et al 1990, Majewski and Trebst 1990 or four peptide subunits (Yu et al 1984, Ljungdhal et al 1987, Andrews et al 1990, Purvis et al 1990, Usui and Yu 1991. Cytochrome bc I complexes isolated from mitochondria contain at least nine subunits (Teintze et al 1982).…”

“…Genes encoding the Rieske iron-sulfur protein, cytochrome b and cytochrome c I from four species of photosynthetic purple non-sulfur bacteria (Gabellini and Sebald 1986, Davidson and Daldal 1987a, Davidson and Daldal 1987b, Verbist et al 1989, Majewski and Trebst 1990, Yun et al 1990), the aerobic bacterium Paracoccus denitrificans (Kurowski and Ludwig 1987), the nitrogen fixing bacterium Bradyrhizobium japonicum (Thony-Meyer et al 1989), and, for the cytochrome b6f complex, from the cyanobacterium Nostoc PCC 7906 (Kallas et al 1988b), have been cloned and sequenced.…”

Identification, cDNA Sequence and Deduced Amino Acid Sequence of the Mitochondrial Rieske Iron‐Sulfur Protein from the Green Alga Chlamydomonas reinhardtii

“…The thick, black bar indicates the sequence which is believed to interact with cytochrome b, based on similarity with the sequence of the bovine enzyme (Gonzhlez-Halphen et al, 1991); two thinner black bars indicate the strictly conserved hexapeptides that are believed to provide the ligands for the iron-sulfur cluster; +, the positively charged residue in the sequence of the mitochondrial Rieske protein from photosynthetic organisms. Sequences of the Rieske proteins from potato mitochondrion (Emmerniann et al, 1994), tobacco mitochondrion clone RISP2 (Huang et al, 1991, bovine mitochondrion (Schagger et al, 1987;Brandt et al, 1993), human mitochondrion (Nishimiki et al, 1990) Neurosporu crassa mitochondrion (Harnisch et al, 1985), R. rubrum (Majewski and Trebst, 1990) and C. reinhardtii chloroplast (de Vitry, 1994).…”