Effect of redox stressors on wild type Mak16, its C38A variant and Rpf1 binding. Credit: Proceedings of the National Academy of Sciences (2025). DOI: 10.1073/pnas.2513844122

A single iron-sulfur building block directly determines whether ribosomes—the protein factories of our cells—work smoothly or not. This is the conclusion of a recent research project led by the RPTU Kaiserslautern-Landau. The findings significantly expand our understanding of the roles of metal ions in protein production and have been published in the Proceedings of the National Academy of Sciences.

Metal ions are important building blocks of life: Iron ions, for example, play a central role in so-called iron-sulfur clusters in proteins, i.e., protein molecules that are involved in various essential biological processes. These include metabolic pathways such as the mitochondrial respiratory chain and the citric acid cycle. Researchers at RPTU have now been able to show that a tiny metal building block is also crucial for protein production itself.

PD Dr. Daili Netz, from the Department of Chemistry at RPTU, and doctoral student Nadine Duppe took a close look at the protein Mak16. It is important to note that Mak16 plays a key role in the production of ribosomes, i.e., the “protein factories” in our cells.

Netz’s team found that Mak16—in the course of ribosome assembly—is only stable and works correctly with its important partner protein Rpf1 if it contains the iron-sulfur building block [4Fe-4S]. The [4Fe-4S] cluster has a structure in which four iron and four sulfur ions are arranged in an approximately cubic pattern, alternating at the corners of the “cube.” If this building block is missing, then—to put it simply—ribosome production breaks down and the cell can no longer produce new proteins.

If the cluster is missing, the ribosomes cannot be assembled properly

“Mak16 carries an iron-sulfur building block in a pocket in the protein,” says Netz, explaining the detailed structure of the compound. “This pocket consists of four amino acids, the cysteine residues, which hold the cluster in place and help it bind stably to the protein.”

To demonstrate how important this cluster is for its interaction with Rpf1, the research team specifically produced Mak16 in two variants: one in its “normal” form with an intact pocket and cluster, and one modified so that the pocket can no longer hold the cluster. Using immunoprecipitation, which can be thought of as a kind of “protein fishing,” the researchers were able to show that only Mak16 with an intact pocket and cluster can reliably hold the Rpf1 protein.

“If the cluster is missing, the binding does not work at all, and no complex is formed,” explains Netz—adding, with regard to the fact that ribosomes consist of proteins and ribosomal RNA (rRNA): “We also looked at whether the ribosomes are assembled correctly in yeast cells. And we could see that the production of rRNA and the maturation of ribosomes strongly depends on whether Mak16 carries the cluster. If the cluster is missing, the ribosomes cannot be assembled properly.”

Assembly of Fe/S clusters on Mak16 and efficient rRNA synthesis relies on the ISC and CIA machinery pathways. Credit: Proceedings of the National Academy of Sciences (2025). DOI: 10.1073/pnas.2513844122

Details about the iron-sulfur building block clarified

Under the direction of Professor Antonio Pierik, Department of Chemistry at RPTU, the metallic nature of the cluster was demonstrated using electron spin resonance (EPR) spectroscopy, supplemented by Mössbauer analyses by Professor Volker Schünemann, Department of Physics at RPTU, and his doctoral student Lukas Knauer. Mössbauer spectroscopy can be understood as a kind of super-precise iron scanner—because using this highly specialized method, the researchers were able to analyze, among other things, how the iron building blocks are bound in the protein structure.

Pierik explains the background of these investigations: “With EPR spectroscopy, we can see the iron ions because they have unpaired electrons that produce a fingerprint in a magnetic field at very low temperatures. The sulfur ions themselves cannot be seen directly, but they influence the iron ions so strongly that their presence and arrangement can also be detected.”

EPR and Mössbauer analyses show that Mak16 contains a [4Fe-4S] cluster that occurs in two stable states. Netz says, “This enabled us to understand how the metal ions are organized in the protein and how Mak16 can thereby fulfill its tasks in the cell.”

Another finding by the researchers is that the [4Fe-4S] cluster is very sensitive to oxidative stress. If the cluster disintegrates as a result, ribosome production stops. Thus, the cluster acts not only as an essential building block, but also as a sensor that signals to the cell when protein production should be reduced.

“Iron-sulfur clusters control central cellular processes such as metabolism, DNA synthesis and repair, signal transduction, and enzymatic functions, and help cells respond to stress. The fact that a single [4Fe-4S] cluster directly influences ribosome assembly gives us new insights into the mechanisms of protein production, expands our understanding of cell biology, and explains how disruptions in these processes can lead to problems in protein production or cellular stress responses,” says Netz.

More information: Nadine Duppe et al, The function of Mak16 in ribosome biogenesis depends on its [4Fe-4S] cluster, Proceedings of the National Academy of Sciences (2025). DOI: 10.1073/pnas.2513844122

Citation: Iron-sulfur cluster found essential for proper ribosome assembly in cells (2025, November 17) retrieved 17 November 2025 from https://phys.org/news/2025-11-iron-sulfur-cluster-essential-proper.html

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