The c-ring stoichiometry of ATP synthase is adapted to cell physiological requirements of alkaliphilic Bacillus pseudofirmus OF4

Abstract

The c-rings of ATP synthases consist of individual c-subunits, all of which harbor a conserved motif of repetitive glycine residues (GxGxGxG) important for tight transmembrane α-helix packing. The cring stoichiometry determines the number of ions transferred during enzyme operation and has a direct impact on the ion-to-ATP ratio, a cornerstone parameter of cell bioenergetics. In the extreme alkaliphile Bacillus pseudofirmus OF4, the glycine motif is replaced by AxAxAxA. We performed a structural study on two mutants with alanine-to-glycine changes using atomic force microscopy and X-ray crystallography, and found that mutants form smaller c 12 rings compared with the WT c 13 . The molar growth yields of B. pseudofirmus OF4 cells on malate further revealed that the c 12 mutants have a considerably reduced capacity to grow on limiting malate at high pH. Our results demonstrate that the mutant ATP synthases with either c 12 or c 13 can support ATP synthesis, and also underscore the critical importance of an alanine motif with c 13 ring stoichiometry for optimal growth at pH >10. The data indicate a direct connection between the precisely adapted ATP synthase c-ring stoichiometry and its ionto-ATP ratio on cell physiology, and also demonstrate the bioenergetic challenges and evolutionary adaptation strategies of extremophiles.

Cardiolipin Is Dispensable for Oxidative Phosphorylation and Non-fermentative Growth of Alkaliphilic Bacillus pseudofirmus OF4

“…Further studies in this model system will explore the possibility that several such mechanisms make contributions in this extremophile. In keeping with early proposals of Williams (64,65), several adaptations in the OXPHOS machinery itself have already been shown to be necessary for the OXPHOS capacity displayed by B. pseudofirmus OF4 (38,66,67).…”

A New Type of Na+-Driven ATP Synthase Membrane Rotor with a Two-Carboxylate Ion-Coupling Motif

“…All these organisms thus use the SMF to sustain ATP production, but a phylogenetic relationship among them is not apparent. A more evident similarity, besides the near-identical amino acid composition of the Na + -binding sites, is a glycine-rich motif in the N-terminal helix of the c-subunit (Figure S1), which has been shown to have a strong influence on the stoichiometry of the c-ring [14],[37],[38]. It would be therefore tempting to hypothesize that the 11-hairpin architecture might reflect a universal feature of all Na + -driven c-rings in F-type ATP synthases.…”

Structure of a bacterial ATP synthase

“…The noncatalytic a subunits each bind to a magnesium ion (Mg 2+ ) and a nucleotide, while the catalytic b subunits can adopt different conformations and bind to Mg-ADP (bDP), Mg-ATP (bTP), or remain empty (bE). Crystal structures of bacterial F1-ATPases and c-rings from the FO regions of several species have been determined [16][17][18][19][20][21][22][23][24] . Structures of intact ATP synthases from E. coli have been determined to overall resolutions of 6 to 7 Å by cryo-EM, with the FO region showing lower quality than the rest of the maps, presumably due to conformational flexibility 25 .…”