Authors
Lívia S Mészáros, Pierre Ceccaldi, Marco Lorenzi, Holly J Redman, Emanuel Pfitzner, Joachim Heberle, Moritz Senger, Sven T Stripp, Gustav Berggren
Publication date
2020
Journal
Chemical science
Volume
11
Issue
18
Pages
4608-4617
Publisher
Royal Society of Chemistry
Description
Hydrogenases are among the fastest H2 evolving catalysts known to date and have been extensively studied under in vitro conditions. Here, we report the first mechanistic investigation of an [FeFe]-hydrogenase under whole-cell conditions. Functional [FeFe]-hydrogenase from the green alga Chlamydomonas reinhardtii is generated in genetically modified Escherichia coli cells by addition of a synthetic cofactor to the growth medium. The assembly and reactivity of the resulting semi-synthetic enzyme was monitored using whole-cell electron paramagnetic resonance and Fourier-transform Infrared difference spectroscopy as well as scattering scanning near-field optical microscopy. Through a combination of gas treatments, pH titrations, and isotope editing we were able to corroborate the formation of a number of proposed catalytic intermediates in living cells, supporting their physiological relevance. Moreover, a …
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