Oxygen detoxification in anaerobic bacteria : functional genomics and bioenergetics of oxygen reductases – O2StressAnaerob

The single and double deletion mutants of the two membrane-bound oxygen-reductase are available in our laboratory. we are studying the effects of these genes deletion on the physiology of the bacterium and more specially on its capability to growth in the presence of molecular oxygen. Our methodology includes global approaches (transcriptomics) as well as enzymatic activities and cells energization measurements. Structural information on the various components of these systems will be obtained through NMR and cristallographic analyses.

The results clearly point out the extreme resistance of the sulfate reducing bacterium Desulfovibrio vulgaris Hildenborough towards oxygen. The absence of the membrane bound oxygen reductases induces an increase of the sensitivity to oxygen, clearly showing the involvement of these enzymes in oxygen detoxication. The importance of each enzymes in this process becomes now clearer and an electronic link between the periplasmic hydrogen oxidation and the membrane-bound oxygen reduction has been showed for the first time in these anaerobic bacteria.

Project is in progress; perspectives will be defined at the end.

Publication : Ramel, F., Amrani, A., Pieulle, L., Lamrabet, O., Voordouw, G., Seddiki, N., Brèthes, D., Company, M., Dolla, A. and Brasseur, G. (2013) Membrane-bound oxygen reductases of the anaerobic sulfate-reducing Desulfovibrio vulgaris Hildenborough: roles in oxygen defense and electron link with the periplasmic hydrogen oxidation. Microbiology, 159, 2663-2673.
This paper describes the strains sensitivity to oxygen as well as oxygen reductases activities.

Oxygen is the most powerful compound used by aerobic forms of life through complex respiratory chains that all include at least one membrane-bound terminal oxygen reductase. Surprisingly, the increasing number of sequenced genomes revealed the presence of genes encoding these terminal oxidases not only in aerobic but also in a large number of strictly anaerobic microorganisms. While the function of terminal oxidases is easily understandable in aerobes, occurrence of such enzyme in strict anaerobes is more intriguing. Data suggest that a lot of anaerobic microorganisms, although unable to use oxygen as terminal electron acceptor for growth, have developed defence strategies to protect themselves against oxygen damages and exhibit thus aerotolerance. Aerotolerance studies are of great interest to determine new systems involved in oxygen and reactive oxygen species (ROS) detoxification. Desulfovibrio species are one of the most studied microorganisms concerning aerotolerance mechanisms. Indeed, more and more ecological studies show that, although anaerobes, these microorganisms are able to deal with the temporary presence of oxygen in their natural biotopes and even reduce it. In Desulfovibrio species, three systems that differ from their cellular location have been proposed to account for oxygen reduction: a periplasmic system involving both hydrogenases and cytochromes, a cytoplasmic system involving a flavodiiron protein (rubredoxin oxygen oxidoreductase) as terminal oxygen reductase and two membrane-bound systems namely a quinol-bd oxidase and a cytochrome c oxidase belonging to the heme-copper superfamily with unique cofactor content (two c-type hemes in subunit II and the absence of hemes a in subunit I) making it of cc(o/b)o3 type. Only a few data are available concerning the characterization of the membrane-bound oxygen reductases and their physiological function in anaerobic organisms has never been stated which make this project especially interesting and innovative.
The main goal of this project is thus to get a better understanding of the physiological role of these membrane-bound terminal oxidases in anaerobic organisms using Desulfovibrio vulgaris Hildenborough (DvH) as model. For this purpose, the single and double deletion mutants of these two membrane-bound enzymes available in our laboratory represent an important tool to decipher their respective functional role in the cell. In addition, we also possess the deletion mutant strains corresponding to the two soluble periplasmic and cytoplasmic oxygen reduction systems as well as a triple mutant deleted of the membrane and cytoplasmic oxygen reduction systems. The functional genomics approach we propose to develop will allow us to get the molecular and functional characterization of these membrane-bound oxygen reductases in Desulfovibrio as well as their influence in the energy and carbon metabolisms of the bacterium. It will also provide extensive and new information on the respective role and predominance of each system in oxygen detoxification in anaerobic microorganisms and their evolutionary links with regards to the presence of O2 in atmosphere.