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Abstract Hydrogenase enzymes produce H2gas, which can be a potential source of alternative energy. Inspired by the [NiFe] hydrogenases, we report the construction of a de novo‐designed artificial hydrogenase (ArH). The ArH is a dimeric coiled coil where two cysteine (Cys) residues are introduced at tandema/dpositions of a heptad to create a tetrathiolato Ni binding site. Spectroscopic studies show that Ni binding significantly stabilizes the peptide producing electronic transitions characteristic of Ni‐thiolate proteins. The ArH produces H2photocatalytically, demonstrating a bell‐shaped pH‐dependence on activity. Fluorescence lifetimes and transient absorption spectroscopic studies are undertaken to elucidate the nature of pH‐dependence, and to monitor the reaction kinetics of the photochemical processes. pH titrations are employed to determine the role of protonated Cys on reactivity. Through combining these results, a fine balance is found between solution acidity and the electron transfer steps. This balance is critical to maximize the production of NiI‐peptide and protonation of the NiII−H−intermediate (Ni−R) by a Cys (pKa≈6.4) to produce H2.
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Design of a minimal di-nickel hydrogenase peptide
Ancestral metabolic processes involve the reversible oxidation of molecular hydrogen by hydrogenase. Extant hydrogenase enzymes are complex, comprising hundreds of amino acids and multiple cofactors. We designed a 13–amino acid nickel-binding peptide capable of robustly producing molecular hydrogen from protons under a wide variety of conditions. The peptide forms a di-nickel cluster structurally analogous to a Ni-Fe cluster in [NiFe] hydrogenase and the Ni-Ni cluster in acetyl-CoA synthase, two ancient, extant proteins central to metabolism. These experimental results demonstrate that modern enzymes, despite their enormous complexity, likely evolved from simple peptide precursors on early Earth.
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- Award ID(s):
- 2025200
- PAR ID:
- 10493981
- Editor(s):
- Thorp, Holden
- Publisher / Repository:
- Science Advances
- Date Published:
- Journal Name:
- Science Advances
- Edition / Version:
- 1
- Volume:
- 9
- Issue:
- 10
- ISSN:
- 2375-2548
- Subject(s) / Keyword(s):
- Enzyme Hydrogenase nickel acetyl-CoA synthase nickelback
- Format(s):
- Medium: X Other: pdf
- Sponsoring Org:
- National Science Foundation
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Accepted Manuscript1.0
- Journal Article:
- https://doi.org/10.1126/sciadv.abq1990
