Intra- and Inter-Complex Cross-Linking of Subunits in the Quinol Oxidase Super-Complex from Thermophilic Bacillus PS3

Abstract

Gram-positive thermophilic Bacilli contain quinol-cytochrome c reductase and cytochrome c oxidase as two major respiratory complexes of the electron transfer chain, and these enzymes can be extracted with mild detergents as an associated quinol oxidase super-complex. The reductase is composed of three subunits; cytochrome b6, cytochrome c1, and FeS protein, whereas cytochrome c oxidase consists of four subunits numbered 1 through 4. In order to clarify the interactions between the subunits, the super-complex isolated from Bacillus PS3 was cross-linked with three bifunctional cross-linkers; disuccinimidyl tartrate, 3,3'-dithiobis(succinimidylpropionate), and ethylene glycolbis(sulfosuccinimidylsuccinate). The most prominent cross-linking was observed for the combination of subunit 1 plus 2 in cytochrome c oxidase, and for that of cytochrome b6 plus cytochrome c1 in the reductase. In addition to these intra-complex cross-linkings, inter-complex linking was observed for the combination of cytochrome b6 plus subunit 1 with ethylene glycolbis(sulfosuccinimidylsuccinate), and for the combinations of cytochrome b6 plus subunit 1 and cytochrome b6 plus subunit 2 with 3,3'-dithiobis(succinimidylpropionate). Incubation in the presence of Triton X-100, which was confirmed to cleave the two enzyme complexes, selectively reduced the inter-complex cross-linking, suggesting that the chemical cross-linking reflect the spatial arrangement of subunits in the super-complex.

Bacillus subtilis CcdA-defective mutants are blocked in a late step of cytochrome c biogenesis

“…This operon encodes menaquinol:cytochrome c oxidoreductase (the bc complex), which contains a diheme cytochrome b (QcrB; 224 residues) and a monoheme cytochrome c (QcrC; 255 residues) subunit. B. subtilis Qcr mutant data (51, 52) combined with results for Bacillus strain PS3 (18,43) and Bacillus stearothermophilus (41) suggest that the labeled 22-and 29-kDa polypeptides correspond to QcrB and QcrC, respectively.…”

Phylogeny of Rieske/cytb Complexes with a Special Focus on the Haloarchaeal Enzymes

“…The presence of this enzyme allows optimizing the energy yield of diverse respiratory metabolisms, due to the efficient proton pumping capacity of the complex. Rieske/cyt b complexes and their functional role in bioenergetic chains are reasonably well characterized in Heliobacteria (Kramer et al 1997; Ducluzeau et al 2008), Bacilli (Yu and Le Brun 1988; Sone and Fujiwara 1991; Liebl et al 1992; Tanaka et al 1996), Actinobacteria (Sone et al 2001, 2003), and Aquificales (Schütz et al 2003). They are extremely well studied in proteobacteria and mitochondria (p.e.…”

Supramolecular organizations in the aerobic respiratory chain of Escherichia coli

“…strain 7 has a terminal oxygen reductase supercomplex resulting from the functional fusion of complexes III and IV, containing cytochromes of the b and a types, and a Rieske-type ironesulfur protein [17,18]. A mitochondrial-like respirasome was identified in Paraccocus denitrificans [19] and supramolecular associations of complexes III and IV were detected in P. denitrificans [19,20], Corynebacterium glutamicum [21], and Bacillus PS3 [22,23].…”