The composition of the Bacillus subtilis aerobic respiratory chain supercomplexes

Abstract

Bacillus subtilis has a bifurcated respiratory chain composed of a cytochrome branch and a quinol oxidase branch. The respiratory complexes of this bacterium have been elucidated mostly by the analysis of the genome and by the isolation of individual complexes. The supramolecular organization of this respiratory chain is not known. In this work, we have analyzed the organization of the supercomplex in membranes isolated from B. subtilis grown in aerobic conditions in a medium with 3 % succinate. We used two different native electrophoretic techniques, clear native electrophoresis (CNE) and blue native electrophoresis (BNE). Using a heme-specific stain and Coomassie blue stain with in-gel activity assays followed by mass spectrometry, we identified the proteins resolved in both the first and second dimensions of the electrophoreses to detect the supercomplexes. We found that complexes b ( 6 ) c and caa ( 3 ) form a very high molecular mass supercomplex with the membrane-bound cytochrome c ( 550 ) and with ATP synthase. Most of the ATP synthase was found as a monomer. Succinate dehydrogenase was identified within a high molecular band between F(0)F(1) and F(1) and together with nitrate reductase. The type-2 NADH dehydrogenase was detected within a low molecular mass band. Finally, the quinol oxidase aa ( 3 ) seems to migrate as an oligomer of high molecular mass.

Respiration control of multicellularity in Bacillus subtilis by a complex of the cytochrome chain with a membrane-embedded histidine kinase

“…We showed that KinB activation depends on cytochromes caa 3 and bc and that KinB is in a complex with multiple components of the respiratory apparatus, including caa 3 and bc. Interestingly, a recent study indicates that these two cytochromes are in a complex with each other (Garcia Montes de Oca et al 2012).…”

Bacterial Electron Transfer Chains Primed by Proteomics

“…One may note that SDH (complex II) never formed part of mitochondrial SCs. The presence of the succinate:nitrate oxidoreductase and bc:aa 3 SCs in B. subtilis was substantiated in another study (García Montes de Oca et al, 2012), where it was shown that also membrane-bound cytochrome c 550 was a component and the electron carrier of the bc:aa 3 SC. Intriguingly, this SC was associated with ATP synthase monomers.…”

“…11B and C) (Farhoud et al, 2005), and it cannot be ruled out a priori that such dimers and multimers could be found in other prokaryotes, especially when equipped with convolute laminar, tubular or highly curved membrane systems. ATP synthase formed part of the bc:aa 3 SC in B. subtilis (García Montes de Oca et al, 2012). For Bacillus pseudofirmus OF4, an SC has been described consisting of cytochrome aa 3 and ATP synthase (Liu et al, 2007).…”

C‐terminal regulatory domain of the ε subunit of FoF1 ATP synthase enhances the ATP‐dependent H+ pumping that is involved in the maintenance of cellular membrane potential in Bacillus subtilis

“…During the growth‐stalling phase of the ∆ rrn8 ε ∆C mutant, a reduction in the membrane potential, which most likely correlates with the decrease in the PMF, was also observed (Figure c). The PMF of B. subtilis grown under aerobic conditions is mainly generated by the respiratory complex that consumes molecular oxygen as the final electron acceptor (Azarkina et al, ; García Montes de Oca et al, ; Lauraeus & Wikström, ; Winstedt & von Wachenfeldt, ), and thus the activity of the respiratory complex was monitored by measuring the dissolved oxygen concentration in the culture medium. When the respiratory complex activity is high, dissolved oxygen in the culture is continuously consumed, whereas a decrease in the activity of the respiratory complex allows the concentration of dissolved oxygen to increase by dissolution of oxygen into the culture from air (Shimada & Tanaka, ).…”

“…These results suggest that the activity of the respiratory complex in both strains was arrested 3.5 hr after inoculation, and the arrest of activity of the respiratory complex was extended in the ∆ rrn8 ε ∆C mutant. Succinate and NADH, which are driving forces of the respiratory complex (Azarkina et al, ; García Montes de Oca et al, ; Lauraeus & Wikström, ; Winstedt & von Wachenfeldt, ), are synthesized via glycolysis and the TCA cycle. Because glucose is the starting substrate for glycolysis and is metabolized via the TCA cycle, we measured the amount glucose remaining in the culture.…”