In Vitro Biosynthesis of the [Fe]-Hydrogenase Cofactor Verifies the Proposed Biosynthetic Precursors.
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ABSTRACT:
SUBMITTER: Schaupp S
PROVIDER: S-EPMC9314073 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:This communication demonstrates the preparation, isolation, and full characterization of superelectrophilic salts based on amidine dications in organic solvent, as their triflate salts. These dications are highly activated toward regiospecific reaction with hydrogen gas under mild conditions in the presence of a metal catalyst (Pd/C), mimicking the behavior of the natural substrate, N(5),N(10)-methenyltetrahydromethanopterin, in the iron-sulfur cluster-free [Fe]-hydrogenase.
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Project description:Nature carefully selects specific metal ions for incorporation into the enzymes that catalyse the chemical reactions necessary for life. Hydrogenases, enzymes that activate molecular H2, exclusively utilize Ni and Fe in [NiFe]-, [FeFe]- and [Fe]-hydrogeanses. However, other transition metals are known to activate or catalyse the production of hydrogen in synthetic systems. Here, we report the development of a biomimetic model complex of [Fe]-hydrogenase that incorporates a Mn, as opposed to a Fe, metal centre. This Mn complex is able to heterolytically cleave H2 as well as catalyse hydrogenation reactions. The incorporation of the model into an apoenzyme of [Fe]-hydrogenase results in a [Mn]-hydrogenase with an enhanced occupancy-normalized activity over an analogous semi-synthetic [Fe]-hydrogenase. These findings demonstrate a non-native metal hydrogenase that shows catalytic functionality and that hydrogenases based on a manganese active site are viable.
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Project description:Assembly of active Fe-hydrogenase in the chloroplasts of the green alga Chlamydomonas reinhardtii requires auxiliary maturases, the S-adenosylmethionine-dependent enzymes HydG and HydE and the GTPase HydF. Genes encoding homologous maturases had been found in the genomes of all eubacteria that contain Fe-hydrogenase genes but not yet in any other eukaryote. By means of proteomic analysis, we identified a homologue of HydG in the hydrogenosomes, mitochondrion-related organelles that produce hydrogen under anaerobiosis by the activity of Fe-hydrogenase, in the pathogenic protist Trichomonas vaginalis. Genes encoding two other components of the Hyd system, HydE and HydF, were found in the T. vaginalis genome database. Overexpression of HydG, HydE, and HydF in trichomonads showed that all three proteins are specifically targeted to the hydrogenosomes, the site of Fe-hydrogenase maturation. The results of Neighbor-Net analyses of sequence similarities are consistent with a common eubacterial ancestor of HydG, HydE, and HydF in T. vaginalis and C. reinhardtii, supporting a monophyletic origin of Fe-hydrogenase maturases in the two eukaryotes. Although Fe-hydrogenases exist in only a few eukaryotes, related Narf proteins with different cellular functions are widely distributed. Thus, we propose that the acquisition of Fe-hydrogenases, together with Hyd maturases, occurred once in eukaryotic evolution, followed by the appearance of Narf through gene duplication of the Fe-hydrogenase gene and subsequent loss of the Hyd proteins in eukaryotes in which Fe-hydrogenase function was lost.
| S-EPMC1398061 | biostudies-literature
Project description:[Fe-Fe]-hydrogenases are enzymes that reversibly catalyze the reaction of protons and electrons to molecular hydrogen, which occurs in anaerobic media. In living systems, [Fe-Fe]-hydrogenases are mostly used for H(2) production. The [Fe-Fe]-hydrogenase H-cluster is the active site, which contains two iron atoms. The latest theoretical investigations1,2 advocate that the structure of di-iron air inhibited species are either Fe(p) (II)-Fe(d) (II)-O-H(-), or Fe(p) (II)-Fe(d) (II)-O-O-H, thus O(2) has to be prevented from binding to Fe(d) in all di-iron subcluster oxidation states in order to retain a catalytically active enzyme. By performing residue mutations on [Fe-Fe]-hydrogenases, we were able to weaken O(2) binding to distal iron (Fe(d)) of Desulfovibrio desulfuricans hydrogenase (DdH). Individual residue deletions were carried out in the 8 A apoenzyme layer radial outward from Fe(d) to determine what residue substitutions should be made to weaken O(2) binding. Residue deletions and substitutions were performed for three di-iron subcluster oxidation states, Fe(p) (II)-Fe(d) (II), Fe(p) (II)-Fe(d) (I), and Fe(p) (I)-Fe(d) (I) of [Fe-Fe]-hydrogenase. Two deletions (DeltaThr(152) and DeltaSer(202)) were found most effective in weakening O(2) binding to Fe(d) in Fe(p) (II)-Fe(d) (I) hydrogenase (DeltaG(QM/MM) = +5.4 kcal/mol). An increase in Gibbs' energy (+2.2 kcal/mol and +4.4 kcal/mol) has also been found for Fe(p) (II)-Fe(d) (II), and Fe(p) (I)-Fe(d) (I) hydrogenase respectively. pi-backdonation considerations for frontier molecular orbital and geometrical analysis corroborate the Gibbs's energy results.
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Project description:The oxidation of H-cluster in gas phase, and in aqueous enzyme phase, has been investigated by means of quantum mechanics (QM) and combined quantum mechanics-molecular mechanics (QM/MM). Several potential reaction pathways (in the above mentioned chemical environments) have been studied, wherein only the aqueous enzyme phase has been found to lead to an inhibited hydroxylated cluster. Specifically, the inhibitory process occurs at the distal iron (Fe(d)) of the catalytic H-cluster (which is also the atom involved in H(2) synthesis). The processes involved in the H-cluster oxidative pathways are O(2) binding, e(-) transfer, protonation, and H(2)O removal.We found that oxygen binding is non-spontaneous in gas phase, and spontaneous for aqueous enzyme phase where both Fe atoms have oxidation state II; however, it is spontaneous for the partially oxidized and reduced clusters in both phases. Hence, in the protein environment the hydroxylated H-cluster is obtained by means of completely exergonic reaction pathway starting with proton transfer.A unifying endeavor has been carried out for the purpose of understanding the thermodynamic results vis-à-vis several other performed electronic structural methods, such as frontier molecular orbitals (FMO), natural bond orbital partial charges (NBO), and H-cluster geometrical analysis. An interesting result of the FMO examination (for gas phase) is that an e(-) is transferred to LUMO(alpha) rather than to SOMO(beta), which is unexpected because SOMO(beta) usually resides in a lower energy rather than LUMO(alpha) for open-shell clusters.
| S-EPMC2613324 | biostudies-literature
Project description:[FeFe] hydrogenases carry out the redox interconversion of protons and molecular hydrogen (2H+ + 2e- ⇌ H2) at a complex Fe-S active site known as the H-cluster. The H-cluster consists of a [4Fe-4S] subcluster, denoted here as [4Fe]H, linked via a cysteine sulfur to an interesting organometallic [2Fe]H subcluster thought to be the subsite where the catalysis occurs. This [2Fe]H subcluster consists of two Fe atoms, linked with a bridging CO and a bridging SCH2NHCH2S azadithiolate (adt), with additional terminal CO and CN ligands bound to each Fe. Synthesizing such a complex organometallic unit is a fascinating problem in biochemistry, complicated by the toxic nature of both the CO and CN- species and the relative fragility of the azadithiolate bridge. It has been known for a number of years that this complex biosynthesis is carried out by a set of three essential Fe-S proteins, HydE, HydF, and HydG. HydF is a GTPase, while HydE and HydG are both members of the large family of radical S-adenosylmethionine (rSAM) enzymes. In this perspective we describe the history of research and discovery concerning these three Fe-S "maturase" proteins and describe recent evidence for a sequential biosynthetic pathway beginning with the synthesis of a mononuclear organometallic [Fe(ii)(CO)2CN(cysteine)] complex by the rSAM enzyme HydG and its subsequent activation by the second rSAM enzyme HydE to form a highly reactive Fe(i)(CO)2(CN)S species. In our model a pair of these Fe(i)(CO)2(CN)S units condense to form the [Fe(CO)2(CN)S]2 diamond core of the [2Fe]H cluster, requiring only the installation of the central CH2NHCH2 portion of the azadithiolate bridge, whose atoms are all sourced from the amino acid serine. This final step likely occurs with an interplay of HydE and HydF, the details of which yet remain to be elucidated.
| S-EPMC8162317 | biostudies-literature