Immobilization of the hyperthermophilic hydrogenase from Aquifex aeolicus bacterium onto gold and carbon nanotube electrodes for efficient H2 oxidation

Abstract

The electrochemistry of membrane-bound [NiFe] hydrogenase I ([NiFe]-hase I) from the hyperthermophilic bacterium Aquifex aeolicus was investigated at gold and graphite electrodes. Direct and mediated H(2) oxidation were proved to be efficient in a temperature range of 25-70 degrees C, describing a potential window for H(2) oxidation similar to that of O(2)-tolerant hydrogenases. Search for enhancement of current densities and enzyme stability was achieved by the use of carbon nanotube coatings. We report high catalytic currents for H(2) oxidation up to 1 mA cm(-2), 10 times higher than at the bare electrode. Interestingly, high stability of the direct catalytic process was observed when encapsulating A. aeolicus [NiFe]-hase I into a carboxylic functionalized single walled carbon nanotube network. This suggests a peculiar interaction between the enzyme and the electrode material. The parameters that governed the orientation of the enzyme before electron transfer were thus investigated using self-assembled-monolayer gold electrodes. No control of the orientation by the charge or the hydrophobicity of the interface was demonstrated. This behavior was explained on the basis of a structural comparison between A. aeolicus [NiFe]-hase I and Desulfovibrio fructosovorans [NiFe] hydrogenase, which revealed the absence of acidic residues and an additional loop in the environment of the [4Fe-4S] distal cluster in A. aeolicus [NiFe]-hase I. Finally, the effect of inhibitors on the direct oxidation of H(2) by A. aeolicus [NiFe]-hase I encapsulated in a single walled carbon nanotube network was investigated. No inhibition by CO and tolerance toward O(2) were observed. Discussion of the reasons for such tolerance was undertaken on the basis of structural comparison with hydrogenases from aerobic bacteria.

Biohydrogen for a New Generation of H2/O2 Biofuel Cells: A Sustainable Energy Perspective

“…[105] All the O 2 -tolerant Hases share the same phenotypes in addition to O 2 tolerance: electron transfer from the [NiFe] active site to a membrane-bound diheme cytochrome b through a conductive line of three FeS clusters (electrons are then transferred via a quinone pool to a bc cytochrome then to a cytochrome oxidase) [96] ( Figure 5 A), CO-insensitivity (in contrast to O 2 -sensitive Hases), and higher redox potentials of the FeS clusters than the clusters in O 2 -sensitive Hases (Table 2). [77,78,[106][107][108][109][110][111][112] In addition, because it is purified from a hyperthermophilic bacterium, Aa MbH is able to oxidize H 2 over a temperature range from 25 to 85 8C. [106] The O 2 -tolerant Hases are also much worse H 2 producers than O 2 -sensitive Hases.…”

“…[77,78,[106][107][108][109][110][111][112] In addition, because it is purified from a hyperthermophilic bacterium, Aa MbH is able to oxidize H 2 over a temperature range from 25 to 85 8C. [106] The O 2 -tolerant Hases are also much worse H 2 producers than O 2 -sensitive Hases. [106,113] This is accompanied by an overpotential requirement for H 2 oxidation.…”

Characterization of a unique [FeS] cluster in the electron transfer chain of the oxygen tolerant [NiFe] hydrogenase from Aquifex aeolicus

“…One of the major limitations of hydrogenases, however, is their sensitivity towards oxygen. Recently, the discovery of hydrogenases that retain catalytic activity in oxygenic environments has potentially opened new applications as "green" vanguard catalysts, in particular as electrocatalysts on electrodes for biofuel cells (3,4).Aquifex aeolicus is a hyperthermophilic Knallgas bacterium with optimum growth temperature of 85°C (5). This microorganism harbors three distinct [NiFe] hydrogenases, among which Hase I is located in the aerobic respiration pathway and attached to the membrane via a diheme cytb (6).…”

“…One of the major limitations of hydrogenases, however, is their sensitivity towards oxygen. Recently, the discovery of hydrogenases that retain catalytic activity in oxygenic environments has potentially opened new applications as "green" vanguard catalysts, in particular as electrocatalysts on electrodes for biofuel cells (3,4).…”

“…Hase I consists of two subunits; the large subunit contains the hetero-bimetallic nickel-iron site and the small subunit the electron transfer cofactors, namely iron-sulfur clusters (6). Based on spectroelectrochemical studies, this enzyme exhibits enhanced thermostability and tolerance for inhibitors (e.g., O 2 and CO) (4,7).…”