The catalytic center of [FeFe] hydrogenases, in which the molecular hydrogen oxidation or synthesis takes place, is called the H-cluster. It is a highly complex metallo-organic structure that is composed of a canonical [4Fe4S] cubane-cluster that, through a cysteine residue, binds a unique [FeFe] subcluster, which is coordinated with three CO, two CN⁻, and one unusual azadithiolate bridging ligand.

Jan Tachezy Research Group

Significance

Trichomonads, represented by the highly prevalent sexually transmitted human parasite Trichomonas vaginalis, are anaerobic eukaryotes with hydrogenosomes in the place of the standard mitochondria. Hydrogenosomes form indispensable FeS-clusters, synthesize ATP, and release molecular hydrogen as a waste product. Hydrogen formation is catalyzed by [FeFe] hydrogenase, the hallmark enzyme of all hydrogenosomes found in various eukaryotic anaerobes. Eukaryotic hydrogenases were originally thought to be exclusively localized within organelles, but today few eukaryotic anaerobes are known that possess hydrogenase in their cytosol. We identified a thus-far unknown hydrogenase in T. vaginalis cytosol that cannot use ferredoxin as a redox partner but can use cytochrome b5 as an electron acceptor. Trichomonads overexpressing the cytosolic hydrogenase, while maintaining the carbon flux through hydrogenosomes, show decreased excretion of hydrogen and increased excretion of methylated alcohols, suggesting that the cytosolic hydrogenase uses the hydrogen gas as a source of reducing power for the reactions occurring in the cytoplasm and thus accounts for the overall redox balance. This is the first evidence of hydrogen uptake in a eukaryote, although further work is needed to confirm it. Assembly of the catalytic center of [FeFe] hydrogenases (H-cluster) requires the activity of three dedicated maturases, and these proteins in T. vaginalis are exclusively localized in hydrogenosomes, where they participate in the maturation of organellar hydrogenases. Despite the different subcellular localization of cytosolic hydrogenase and maturases, the H-cluster is present in the cytosolic enzyme, suggesting the existence of an alternative mechanism of H-cluster assembly.

Smutna, T. ; Dohnalkova, A. ; Sutak, R. ; Narayanasamy, R.K.; Tachezy, J.^, and Hrdy, I.: A cytosolic ferredoxin-independent hydrogenase possibly mediates hydrogen uptake in Trichomonas vaginalis, Current Biol. 2022, 32, R49-R51, https://doi.org/10.1016/j.cub.2021.10.050

Cryo-EM structures of CspA27 inside the exit tunnel

1. Structure and nascent chain contacts in the exit tunnel for CspA27-1.
2. Structure and nascent chain contacts in the exit tunnel for CspA27-2.
3. Structure and nascent chain contacts in the exit tunnel for CspA27-3.

Marina V. Rodnina Research Group

Significance

Cellular proteins begin to fold as they emerge from the ribosome. The folding landscape of nascent chains is not only shaped by their amino acid sequence but also by the interactions with the ribosome. Here, we combine biophysical methods with cryo-EM structure determination to show that folding of a β-barrel protein begins with formation of a dynamic α-helix inside the ribosome. As the growing peptide reaches the end of the tunnel, the N-terminal part of the nascent chain refolds to a β-hairpin structure that remains dynamic until its release from the ribosome. Contacts with the ribosome and structure of the peptidyl transferase center depend on nascent chain conformation. These results indicate that proteins may start out as α-helices inside the tunnel and switch into their native folds only as they emerge from the ribosome. Moreover, the correlation of nascent chain conformations with reorientation of key residues of the ribosomal peptidyl-transferase center suggest that protein folding could modulate ribosome activity.

Agirrezabala, X., Samatova, E., Macher, M., Liutkute, M., Maiti, M., Gil-Carton, D., Novacek, J., Valle. M., and Rodnina, M.V.: EMBO J. (2022) e109175, https://doi.org/10.15252/embj.2021109175