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The Rnf complex is a Na⁺ coupled respiratory enzyme in a fermenting bacterium, Thermotoga maritima.

ABSTRACT: rnf genes are widespread in bacteria and biochemical and genetic data are in line with the hypothesis that they encode a membrane-bound enzyme that oxidizes reduced ferredoxin and reduces NAD and vice versa, coupled to ion transport across the cytoplasmic membrane. The Rnf complex is of critical importance in many bacteria for energy conservation but also for reverse electron transport to drive ferredoxin reduction. However, the enzyme has never been purified and thus, ion transport could not be demonstrated yet. Here, we have purified the Rnf complex from the anaerobic, fermenting thermophilic bacterium Thermotoga maritima and show that is a primary Na⁺ pump. These studies provide the proof that the Rnf complex is indeed an ion (Na⁺) translocating, respiratory enzyme. Together with a Na⁺-F₁F_O ATP synthase it builds a simple, two-limb respiratory chain in T. maritima. The physiological role of electron transport phosphorylation in a fermenting bacterium is discussed.

SUBMITTER: Kuhns M

PROVIDER: S-EPMC7414866 | biostudies-literature | 2020 Aug

REPOSITORIES: biostudies-literature

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The Rnf complex is a Na<sup>+</sup> coupled respiratory enzyme in a fermenting bacterium, Thermotoga maritima.

Kuhns Martin M Trifunović Dragan D Huber Harald H Müller Volker V

Communications biology 20200807 1

rnf genes are widespread in bacteria and biochemical and genetic data are in line with the hypothesis that they encode a membrane-bound enzyme that oxidizes reduced ferredoxin and reduces NAD and vice versa, coupled to ion transport across the cytoplasmic membrane. The Rnf complex is of critical importance in many bacteria for energy conservation but also for reverse electron transport to drive ferredoxin reduction. However, the enzyme has never been purified and thus, ion transport could not be ...[more]

PMID: 32770029

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Project description:UnlabelledRegulation of coenzyme A (CoA) biosynthesis in bacteria and eukaryotes occurs through feedback inhibition targeting type I and type II pantothenate kinase (PanK), respectively. In contrast, the activity of type III PanK is not affected by CoA. As the hyperthermophilic bacterium Thermotoga maritima harbors only a single type III PanK (Tm-PanK), here we examined the mechanisms that regulate CoA biosynthesis in this organism. We first examined the enzyme responsible for the ketopantoate reductase (KPR) reaction, which is the target of feedback inhibition in archaea. A classical KPR homolog was not present on the T. maritima genome, but we found a homolog (TM0550) of the ketol-acid reductoisomerase (KARI) from Corynebacterium glutamicum, which exhibits KPR activity. The purified TM0550 protein displayed both KPR and KARI activities and was designated Tm-KPR/KARI. When T. maritima cell extract was subjected to anion-exchange chromatography, the fractions containing high levels of KPR activity also displayed positive signals in a Western blot analysis using polyclonal anti-TM0550 protein antisera, strongly suggesting that Tm-KPR/KARI was the major source of KPR activity in the organism. The KPR activity of Tm-KPR/KARI was not inhibited in the presence of CoA. We thus examined the properties of Tm-PanK and the pantothenate synthetase (Tm-PS) of this organism. Tm-PS was not affected by CoA. Surprisingly however, Tm-PanK was inhibited by CoA, with almost complete inhibition in the presence of 400 μM CoA. Our results suggest that CoA biosynthesis in T. maritima is regulated by feedback inhibition targeting PanK, although Tm-PanK is a type III enzyme.ImportanceBacteria and eukaryotes regulate the biosynthesis of coenzyme A (CoA) by feedback inhibition targeting type I or type II pantothenate kinase (PanK). The hyperthermophilic bacterium Thermotoga maritima harbors a single type III PanK (Tm-PanK), previously considered to be unaffected by CoA. By examining the properties of three enzymes involved in CoA biosynthesis in this organism, we found that Tm-PanK, although a type III enzyme, is inhibited by CoA. The results provide a feasible explanation of how CoA biosynthesis is regulated in T. maritima, which may also apply for other bacteria that harbor only type III PanK enzymes.

Dataset Information

The Rnf complex is a Na⁺ coupled respiratory enzyme in a fermenting bacterium, Thermotoga maritima.

Publications

The Rnf complex is a Na<sup>+</sup> coupled respiratory enzyme in a fermenting bacterium, Thermotoga maritima.

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Dataset Information

The Rnf complex is a Na+ coupled respiratory enzyme in a fermenting bacterium, Thermotoga maritima.

Publications

The Rnf complex is a Na<sup>+</sup> coupled respiratory enzyme in a fermenting bacterium, Thermotoga maritima.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

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The Rnf complex is a Na⁺ coupled respiratory enzyme in a fermenting bacterium, Thermotoga maritima.