Unifying Activity, Structure, and Spectroscopy of [NiFe] Hydrogenases: Combining Techniques To Clarify Mechanistic Understanding

Abstract

Achieving a unified understanding of the mechanism of a multi-center redox enzyme such as [NiFe] hydrogenase is complicated by difficulties in reconciling information obtained using different techniques, and on samples in different physical forms. Measurements of the activity of the enzyme, and of factors which perturb activity, are generally carried out using biochemical

The crystalline state as a dynamic system: IR microspectroscopy under electrochemical control for a [NiFe] hydrogenase

“…Computational modelling studies of the active site suggest that deprotonation of a terminal cysteine thiol ligand to Ni causes ν CO to shift to lower energy by ca 30 cm -1 . 73 This shift upon deprotonation matches the difference in ν CO observed between Ni a -L I and Ni a -L II/III for a range of [NiFe] hydrogenases, 22,72 leading us to postulate that a terminal cysteine thiol is not present in either of the Ni a -L II or Ni a -L III sub-states. The high Hyd1 concentration (8 mM) within single crystals allows us to test this hypothesis further through direct observation of the S-H stretching region (ca 2450 -2600 cm -1 ).…”

“…In addition to providing evidence of Ni a -C/Ni a -L tautomerisation in the crystalline state, it is clear from Figure 5 that the relative proportions of each Ni a -L sub-state also vary with pH, consistent with the behaviour of electrode-adsorbed Hyd1 (Figure S14) where the population of Ni a -L II remains roughly constant above pH 6. 71,72 The mechanistic role of the Ni a -L sub-states as sequential intermediates in proton transfer to/from the [NiFe] active site has been demonstrated in photo-triggered potential jump measurements on Soluble Hydrogenase 1 (SH1) from P. furiosus, 36 and cryogenic photolysis of the [NiFe] hydrogenase from D. vulgaris Miyazaki F. 33,34 The most common representation of 'Ni a -L' invokes protonation of a terminal cysteine-S ligand to Ni at the active site. Evidence of cysteine-S protonation in the Ni a -L I sub-state has been reported in the D. vulgaris Miyazaki F [NiFe] hydrogenase, where H/D labelling suggested the presence of an S-H stretching vibration in Ni a -L I .…”

Bio-inspired, Multifunctional Metal–Thiolate Motif: From Electron Transfer to Sulfur Reactivity and Small-Molecule Activation

“…During the catalytic cycle of the [NiFe]-hydrogenase, the Ni ion is solely responsible of the redox activity, switching between the I, II and III states, but also participates to substrate activation through the formation of bridged hydrides. 44,45 In the case of the [FeFe]-hydrogenase, it is proposed that both Fe ions are redox active, switching between the I and II states, but only one of them is responsible for iron-hydride formation. 46 During the last two decades, numerous synthetic models of the active sites of these two hydrogenases have been described, leading to the development of efficient (electro)catalysts for the Hydrogen Evolution Reaction (HER).…”

Large-scale computational discovery and analysis of virus-derived microbial nanocompartments

“…The specific [NiFe] hydrogenases associated with A-domain Encapsulins generally form cytoplasmic soluble heterotetrameric complexes 45,46 and catalyze the reversible interconversion of H2 to two protons and two electrons. 47 The A-domain Encapsulin-associated [NiFe] hydrogenase of P. furiosus has been partially functionally characterized, however, this was done through whole cell measurements and heterologous expression experiments which did not yield any information about the associated A-domain Encapsulin. 48,49 In P. furiosus, this hydrogenase complex is known as sulfhydrogenase I (SHI) referring to its ability to act as a sulfur reductase, oxidizing H2 whilst simultaneously reducing elemental sulfur or polysulfides to hydrogen sulfide (H2S).…”