Proton Transfer in the Catalytic Cycle of [NiFe] Hydrogenases: Insight from Vibrational Spectroscopy

Abstract

Catalysis of H2 production and oxidation reactions is critical in renewable energy systems based around H2 as a clean fuel, but the present reliance on platinum-based catalysts is not sustainable. In nature, H2 is oxidized at minimal overpotential and high turnover frequencies at [NiFe] catalytic sites in hydrogenase enzymes. Although an outline mechanism has been established for the [NiFe] hydrogenases involving heterolytic cleavage of H2 followed by a first and then second transfer of a proton and electron away from the active site, details remain vague concerning how the proton transfers are facilitated by the protein environment close to the active site. Furthermore, although [NiFe] hydrogenases from different organisms or cellular environments share a common active site, they exhibit a broad range of catalytic characteristics indicating the importance of subtle changes in the surrounding protein in controlling their behavior. Here we review recent time-resolved infrared (IR) spectroscopic studies and IR spectroelectrochemical studies carried out in situ during electrocatalytic turnover. Additionally, we re-evaluate the significant body of IR spectroscopic data on hydrogenase active site states determined through more conventional solution studies, in order to highlight mechanistic steps that seem to apply generally across the [NiFe] hydrogenases, as well as steps which so far seem limited to specific groups of these enzymes. This analysis is intended to help focus attention on the key open questions where further work is needed to assess important aspects of proton and electron transfer in the mechanism of [NiFe] hydrogenases.

From Enzymes to Functional Materials—Towards Activation of Small Molecules

“…Although both enzymes reveal reversible oxidation of H 2 ,[ FeFe]-hydrogenases preferably produces H 2 with ar ate of 10 4 molecules H 2 s À1 and with turnoverf requencies of 10 2 -10 4 s À1 .T his rate is significantly faster than observed for [NiFe]-hydrogenases (10 3 s À1 ), [223] which are known to prefer the H 2 oxidation (> 1000 s À1 ). [224,[225][226][227][228][229] The H-cluster ([2Fe] H )i n[ FeFe]-hydrogenases from Clostridium pasteurianum [231] consists of as ulfur-bridgedp roximal (Fe p ) and distali ron (Fe d )w ith an intermetallic distance of 2.6 coordinated by CO and CN À ligands, as well as an additional bridging CO ligand. [232,233] The dithiol bridgehead contains an amine, which can bend over the distal iron.…”

Photo‐Driven Hydrogen Evolution by an Artificial Hydrogenase Utilizing the Biotin‐Streptavidin Technology

“…Such enzymes display exquisite catalytic properties as they operate: i ) without the need of a high over‐potential, ii ) at high reaction rates (1500 – 20,000 turnovers per second), iii ) under physiological conditions at pH 7 and 37 °C in water. To achieve such unrivalled characteristics, hydrogenases are highly evolved, featuring specialized channels to deliver the substrates and extract the product: proton‐, electron‐, and dihydrogen channels ensure that the reaction does not lead to the generation of side‐products . Importantly, natural hydrogenases rely on earth‐abundant metals ( i.e ., Ni and Fe), but require complex maturation machineries to assemble and display modest stabilities under non‐physiological conditions.…”

“…To achieve such unrivalled characteristics, hydrogenases are highly evolved, featuring specialized channels to deliver the substrates and extract the product: proton-, electron-, and dihydrogen channels ensure that the reaction does not lead to the generation of side-products. [12] Importantly, natural hydrogenases rely on earth-abundant metals (i.e., Ni and Fe), but require complex maturation machineries to assemble and display modest stabilities under non-physiological conditions.…”

Exploitation of Feedback in Enzyme‐catalysed Reactions

“…This enzyme displays multiple redox and coordination states and the details of the transitions between states are still the subject of debate. [36] The reaction involves two parts, an activation process from resting states NiÀA (unready) and NiÀB (ready) followed by heterolytic H 2 cleavage involving short-lived active states Ni a -S, Ni a -C, Ni a -R and possibly Ni a -L: (16) Activation involves a lag phase, the length of which depends on the enzyme concentration and can last several hours. [37] These are hallmark features of an autocatalytic process requiring initiation by a few active hydrogenase molecules.…”

“…The [NiFe] hydrogenase has been isolated from organisms including Thiocapsa roseopersicina and the enzyme activity is quantified in vitro in the presence of a redox partner such as benzyl viologen. This enzyme displays multiple redox and coordination states and the details of the transitions between states are still the subject of debate . The reaction involves two parts, an activation process from resting states Ni−A (unready) and Ni−B (ready) followed by heterolytic H 2 cleavage involving short‐lived active states Ni a ‐S, Ni a ‐C, Ni a ‐R and possibly Ni a ‐L:…”