Ubiquinone Biosynthesis over the Entire O ₂ Range: Characterization of a Conserved O ₂ -Independent Pathway

Abstract

Most bacteria can generate ATP by respiratory metabolism, in which electrons are shuttled from reduced substrates to terminal electron acceptors, via quinone molecules like ubiquinone. Dioxygen (O2) is the terminal electron acceptor of aerobic respiration and serves as a co-substrate in the biosynthesis of ubiquinone. Here, we characterize a novel, O2-independent pathway for the biosynthesis of ubiquinone. This pathway relies on three proteins, UbiT (YhbT), UbiU (YhbU), and UbiV (YhbV). UbiT contains an SCP2 lipid-binding domain and is likely an accessory factor of the biosynthetic pathway, while UbiU and UbiV (UbiU-UbiV) are involved in hydroxylation reactions and represent a novel class of O2-independent hydroxylases. We demonstrate that UbiU-UbiV form a heterodimer, wherein each protein binds a 4Fe-4S cluster via conserved cysteines that are essential for activity. The UbiT, -U, and -V proteins are found in alpha-, beta-, and gammaproteobacterial clades, including several human pathogens, supporting the widespread distribution of a previously unrecognized capacity to synthesize ubiquinone in the absence of O2. Together, the O2-dependent and O2-independent ubiquinone biosynthesis pathways contribute to optimizing bacterial metabolism over the entire O2 range. IMPORTANCE In order to colonize environments with large O2 gradients or fluctuating O2 levels, bacteria have developed metabolic responses that remain incompletely understood. Such adaptations have been recently linked to antibiotic resistance, virulence, and the capacity to develop in complex ecosystems like the microbiota. Here, we identify a novel pathway for the biosynthesis of ubiquinone, a molecule with a key role in cellular bioenergetics. We link three uncharacterized genes of Escherichia coli to this pathway and show that the pathway functions independently from O2. In contrast, the long-described pathway for ubiquinone biosynthesis requires O2 as a substrate. In fact, we find that many proteobacteria are equipped with the O2-dependent and O2-independent pathways, supporting that they are able to synthesize ubiquinone over the entire O2 range. Overall, we propose that the novel O2-independent pathway is part of the metabolic plasticity developed by proteobacteria to face various environmental O2 levels.

The PhoPQ Two-Component System Is the Major Regulator of Cell Surface Properties, Stress Responses and Plant-Derived Substrate Utilisation During Development of Pectobacterium versatile-Host Plant Pathosystems

“…The nrdD gene codes for anaerobic ribonucleoside triphosphate reductase, which is essential in E. coli for deoxyribonucleotide synthesis during strict anaerobic growth ( Garriga et al, 1996 ). ubiQ is required for anaerobic ubiquinone biosynthesis ( Pelosi et al, 2019 ). GrcA restores pyruvate formate-lyase (PFL) activity after oxidative damage to the main PFL subunit ( Wagner et al, 2001 ; Bowman et al, 2019 ) and thus may help the pathogen to maintain the high activity of key glycolysis enzyme at microaerobic conditions.…”

Identification of two fnr genes and characterisation of their role in the anaerobic switch in Sphingopyxis granuli strain TFA

“…Other genes of this category include moeB (SGRAN_0222), an unlinked gene involved in Mo cofactor biosynthesis, most of the components of the cyoABCDEregBA operon, (SGRAN_2997-_2991) encoding a bo3 -type quinol oxidase electron transfer chain and the redox responsive regBA two-component regulatory system 18 . yhbTUV operon (SGRAN_3858-_3860) is one of the most induced operons in anaerobiosis 17 and it is involved in the recently described O 2 -independent ubiquinone biosynthesis pathway, proposed to be renamed ubiTUV 35 , and it is essential for denitrification in Pseudomonas 36 . Althought dRNA-Seq showed that expression of the distal genes of this operon was still maintained to some extent in the Δ narG mutant, the , induction kinetics of ubiU in Δ fnr and in Δ narG mutants (Fig.…”

Advances in bacterial pathways for the biosynthesis of ubiquinone

“…The presence of an iron-sulfur cluster might be another feature common to U32 proteins. Indeed, we showed that UbiU and UbiV each carry a 4Fe-4S cluster ligated by a motif of conserved cysteine residues, which is found in most U32 peptidase family members 46 . Interestingly, the function of RlhA and TrhP depends on these Cys residues and on the genes of the isc operon that catalyze the biogenesis of Fe-S clusters 57,59 .…”

“…Based on the observation that E. coli was able to synthesize UQ under anoxic conditions, the existence of a UQ biosynthesis pathway independent from O2 had long been hypothesized 55 . This pathway remained uncharacterized until 2019, when we identified three genes, ubiT, ubiU and ubiV which are required for UQ biosynthesis under anoxic conditions but are dispensable under oxic conditions 46 we also hypothesized that UbiU and UbiV may participate in the two other hydroxylation steps of the anaerobic UQ pathway, thus substituting for UbiI and UbiH of the O2-dependent pathway (Figure 3A) 46,56 .…”

“…Cells lacking ubiJ show a complete absence of UQ, while ubiK mutants retain ~ 20% UQ compared to wildtype. The UQ deficiency is apparent only when the cells are grown in oxic conditions, suggesting that UbiJ and UbiK do not play important functions for UQ biosynthesis under anoxic conditions 28,45,46 . Purified UbiJ and UbiK interact and form an elongated UbiJ1:UbiK2 complex 28 (Figure 2).…”