Heterodisulfide reductase megacomplexes produced in a Class I CO2-reducing methanogen under Ni-sufficient and Ni-limited conditions
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Philipps-Universität Marburg
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Methanogenic archaea are the major producers of the greenhouse gas methane on Earth. In the methanogenic pathway from CO2 and H2, low-potential electrons for CO2 reduction are formed by a flavin-based electron bifurcation (FBEB) reaction catalyzed by heterodisulfide reductase (Hdr) in complex with Hdr-associating [NiFe] hydrogenase (Mvh). In addition, F420-reducing [NiFe]-hydrogenase (Frh) provides electrons to the methanogenic pathway via an electron carrier F420. It is known that under moderately nickel-limited conditions (Ni2+ ~200 nM), the production of Frh in Methanothermobacter marburgensis is strictly down regulated and the catalytic function is replaced by a coupled reaction of Hmd ([Fe]-hydrogenase) and F420-dependent methylene-tetrahadromethanopterin (methylene-H4MPT) dehydrogenase (Mtd). Proteomic analysis of M. marburgensis in this study indicated that under strictly nickel-limited conditions (Ni2+ < 50 nM), close to the natural environment, Mvh is also not produced, and the production of uncharacterized proteins encoded in a gene cluster are induced [Chapter 2]. We refer to these proteins as F420-dependent electron-donating proteins (ElpABC) and hypothesize that these proteins form a complex with Hdr to donate electrons to the FBEB reaction in this methanogen under strictly nickel-limited conditions. Indeed, the purified Hdr complex contains ElpABC and catalyzed the F420 dependent Hdr reaction. Thus, under strictly nickel-limited conditions often found in nature, Hmd provides all electrons for the reducing metabolism instead of the cytosolic [NiFe]-hydrogenases (Frh and Mvh). Conservation of the elp and hmd genes in the genome of hydrogenotrophic methanogens suggests that this nickel-dependent transition in electron flow occurs in nature. Unexpectedly, the Elp-Hdr complex was purified as a megacomplex with formyl-methanofuran dehydrogenase (Fmd). Cryo-electron microscopy (cryo-EM) revealed the structure of the Elp-Hdr, confirming the electronic interaction between Elp and Hdr although the Fmd module of this megacomplex was dissociated due to the instability of the protein complex. The finding of Elp-Hdr-Fmd megacomplex prompted us to purify the Mvh-Hdr-Fmd complex from M. marburgensis cultivated under nickel-sufficient standard laboratory condition. We successfully purified the Mvh-Hdr-Fmd megacomplex, which was more stable than the Elp-Hdr-Fmd complex. The purified megacomplex catalyzed the H2-dependent electron bifurcation reduction of [13C]-CO2 to [13C]-formyl-methanofuran in the absence of free ferredoxin [Chapter 3]. This result demonstrated that the subunits in the Mvh-Hdr-Fmd megacomplex are electronically linked to transfer electrons for the reduction of CO2. This result also supports the physiological function of the Elp-Hdr-Fmd complex in CO2 reduction using electrons from the reduced form of F420. Most recently, we elucidated the cryo-EM structure of the Mvh-Hdr-Fmd complex from M. marburgensis (unpublished results). The topology of the asymmetric Mvh-Hdr-Fmd complex is distinct from the previously published symmetric Fdh-Hdr-Fmd complex from Methanospirillum hungatei. In the asymmetric Mvh-Hdr-Fmd structure, two FmdABCDFGH dimers form an hourglass-like tetramer, which associates with the Mvh-Hdr dimer. The electron output domain of HdrA and electron input domain of FmdF is connected by an extra polyferredoxin MvhB, which contains twelve [4Fe-4S]-clusters.
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Created: 2024Issued: 2025-11-26Updated: 2024-11-15
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Fachbereich Biologie
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eng
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300
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Nomura, Shunsuke: Heterodisulfide reductase megacomplexes produced in a Class I CO2-reducing methanogen under Ni-sufficient and Ni-limited conditions. : Philipps-Universität Marburg 2025-11-26. DOI: https://doi.org/10.17192/z2024.0485.