Researchers isolate fragile protein complex from heat-loving bacteria

Goethe University scientists have discovered what is probably the oldest enzyme found in cellular respiration. Now, the team has successfully isolated a highly delicate protein complex known as “Rnf” from the heat-loving bacterium called Thermotoga maritima.

The genes encoding the enzyme were already identified about a decade before. But for the first time, the Frankfurt team has successfully isolated the enzyme and thus demonstrated that it is actually formed by microbes and used for producing cellular energy.

The Earth had no oxygen in the first billion years, and life evolved under an anaerobic environment. Perhaps, early bacteria acquired their energy by disintegrating numerous substances through fermentation. But a kind of “oxygen-free respiration” also seemed to exist. This was indicated by studies performed on primordial bacteria that still survive in anaerobic habitats even today.

We already saw ten years ago that there are genes in these microbes that perhaps encode for a primordial respiration enzyme. Since then, we—as well as other groups worldwide—have attempted to prove the existence of this respiratory enzyme and to isolate it. For a long time unsuccessfully because the complex was too fragile and fell apart at each attempt to isolate it from the membrane. We found the fragments, but were unable to piece them together again.”

Volker Müller, Professor, Department of Molecular Microbiology and Bioenergetics, Goethe University

Through perseverance and hard work, Müller’s doctoral researchers Martin Kuhns and Dragan Trifunovic subsequently made a major discovery in two consecutive doctoral theses.

In our desperation, we at some point took a heat-loving bacterium, Thermotoga maritima, which grows at temperatures between 60 °C and 90 °C. Thermotoga also contains Rnf genes, and we hoped that the Rnf enzyme in this bacterium would be a bit more stable. Over the years, we then managed to develop a method for isolating the entire Rnf enzyme from the membrane of these bacteria.”

Dragan Trifunovic, Doctoral Researcher, Goethe University

Trifunovic will soon complete his PhD.

As reported by the team in their latest article, the enzyme complex works somewhat like a pumped-storage power plant, pumping water into a lake higher up and generating electricity through a turbine from the water that flows back down again.

The Rnf enzyme (biochemical name = ferredoxin:NAD-oxidoreductase) transports sodium ions only in the bacterial cell. This enzyme transports the ions from the interior of the cell through the cell membrane to the outside, and while doing so, it creates an electric field.

This electric field is then employed to fuel a cellular “turbine” (ATP synthase): it enables the sodium ions to flow back along the electric field into the interior of the cell and, while doing so, it acquires energy in the form of the cellular energy currency ATP.

The bioenergetic characterization and biochemical evidence of this primordial Rnf enzyme describe how the first life forms created the central energy currency ATP. Evidently, the Rnf enzyme works so well that it is still restricted in several bacteria and in a few archaea today. It is also contained in certain pathogenic bacteria, where the function of the Rnf enzyme is yet to be clearly understood.

Our studies thus radiate far beyond the organism Thermotoga maritima under investigation and are extremely important for bacterial physiology in general.”

Volker Müller, Professor, Department of Molecular Microbiology and Bioenergetics, Goethe University

Müller added that it is now crucial to clearly understand the working of the Rnf enzyme and the kind of role played by the individual parts.

I’m happy to say that we’re well on the way here, since we’re meanwhile able to produce the Rnf enzyme ourselves using genetic engineering methods,” Müller concluded.

Journal reference:

Kuhns, M., et al. (2020) The Rnf complex is a Na+ coupled respiratory enzyme in a fermenting bacterium, Thermotoga maritima. Communications Biology.


The opinions expressed here are the views of the writer and do not necessarily reflect the views and opinions of AZoLifeSciences.
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