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Structural biology: at the heart of the core of bacterial metalloproteins

To be or not to be expressed? Researchers at the CEA-Irig (IBS) are showing with precision how a bacterial metalloprotein controls the expression of certain genes. And all it takes is an electron and a proton!

Published on 13 May 2020

Although living organisms are essentially composed of organic matter, a great number of natural processes depend directly on inorganic factors. Indeed, nearly 40% of proteins require the binding of one or more metal ions (sodium, magnesium, calcium, iron, zinc, copper, etc.) in order to function. These proteins are metalloproteins, and they contain inorganic aggregates that are involved in biosynthetic and metabolic reactions highly important for cellular life. For example, iron-sulfur [Fe-S] aggregates, which are ubiquitous in animals, plants and bacteria, are essential for enzyme catalysis, electron transfer (respiration, photosynthesis) and the regulation of gene expression.

Researchers at the CEA-Irig (IBS) were interested in the bacterial metalloprotein RsrR, which has a [2Fe-2S] center. RsrR is involved in the metabolic regulation of the cell by repressing the expression of certain genes. In early 2019, the scientists determined its crystal structure using X-ray diffraction studies, and their British collaborators showed that it does not bind to DNA when the protein is reduced, i.e. when its [2Fe-2S] center captures an electron. Their recent work has gone further in explaining this phenomenon.

Juan C. Fontecilla-Camps, a researcher at the Irig (IBS), explains their hypothesis: “We assumed, thanks to the crystal structures, that when the protein is reduced, the side chain of a tryptophan amino acid residue turns towards the inside of the protein, preventing it from attaching to the DNA.” To verify this, the scientists attempted to chemically modify tryptophan in the two configurations, either reduced (with an additional negatively charged electron), or oxidized (without this electron). In the former case they did not succeed, validating the hypothesis that the tryptophan is buried inside the RsrR protein (making it no longer accessible). Conversely, the tryptophan was modified in the oxidized configuration, confirming that it is exposed to the solvent.

The researchers also uncovered the behavior of another amino acid, histidine, concomitant to that of tryptophan. “When RsrR is reduced, this histidine moves to the metal center of the protein,” adds Juan C. Fontecilla-Camps. “We have shown using theoretical calculations (in collaboration with the CEA-Irig (DIESE)) that the side chain of this amino acid is then protonated and that, thanks to its positive charge, it is attracted to the [2Fe-2S] center. This histidine then makes way for the tryptophan to bury itself in the protein.” Finally, this hypothesis was confirmed by the crystal structure of an RsrR-DNA complex.

This work shows how effectors as small as an electron and a proton can induce structural changes responsible for an adapted response of the bacteria to its environment!

Interdisciplinary Research Institute of Grenoble (IRIG - CEA/CNRS/Université Grenoble Alpes)
University of East Anglia, Norwich (UK)
​Paul Scherrer Institute (Switzerland)

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