Science Advances 2022

Overview of hcis-PT structure and reaction scheme

(A) Cartoon representation of a single DHDDS-NgBR heterodimer in complex with FPP [Protein Data Bank (PDB) 6Z1N]. DHDDS and NgBR are colored blue and yellow, respectively. Surface representations of the FPP and IPP [placed by superposition with PDB 6W2L] molecules are colored pink and green, respectively. The residue W3, at the DHDDS N terminus, is shown as spheres. (B) Condensation reaction scheme. At the first cycle, the allylic diphosphate primer, FPP (C₁₅, pink), undergoes a condensation with IPP (C₅, green) to produce geranylgeranyl-diphosphate (GGPP) (C₂₀). The cycle repeats with further condensations (14–17) of the allylic diphosphate at S₁, ultimately leading to a final product length of C_85–100.

Yoni Haitin Research Group

Significance

Isoprenoids are synthesized by the prenyltransferase superfamily, which is subdivided according to the product stereoisomerism and length. In short- and medium-chain isoprenoids, product length correlates with active site volume. However, enzymes synthesizing long-chain products and rubber synthases fail to conform to this paradigm, because of an unexpectedly small active site. Here, we focused on the human cis-prenyltransferase complex (hcis-PT), residing at the endoplasmic reticulum membrane and playing a crucial role in protein glycosylation. Crystallographic investigation of hcis-PT along the reaction cycle revealed an outlet for the elongating product. Hydrogen-deuterium exchange mass spectrometry analysis showed that the hydrophobic active site core is flanked by dynamic regions consistent with separate inlet and outlet orifices. Last, using a fluorescence substrate analog, we show that product elongation and membrane association are closely correlated. Together, our results support direct membrane insertion of the elongating isoprenoid during catalysis, uncoupling active site volume from product length.

Giladi, M., Bar-El, ML., Lisnyansky, M., Vankova, P., Ferofontov, A., Melvin, E., Alkaderi, S., Kavan, D., Redko, B., Haimov, E., Wiener, R., Man, P., and Haitin, Y.: Structural basis for long-chain isoprenoid synthesis by cis-prenyltransferases, Sci. Adv. 2022, 7, eabn1171, https://doi.10.1126/sciadv.abn1171

Current Biology 2022

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. & 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