{"id":725,"date":"2017-04-26T11:36:59","date_gmt":"2017-04-26T15:36:59","guid":{"rendered":"http:\/\/sites.williams.edu\/bigchem\/?page_id=725"},"modified":"2017-05-26T10:44:57","modified_gmt":"2017-05-26T14:44:57","slug":"nife-hydrogenase","status":"publish","type":"page","link":"https:\/\/sites.williams.edu\/bigchem\/nife-hydrogenase\/","title":{"rendered":"[NiFe]-Hydrogenase"},"content":{"rendered":"

<\/a>Summary<\/b><\/h1>\n

[NiFe] hydrogenases are bacterial enzymes responsible for the heterolytic cleavage of hydrogen. The protein structure contains a NiFe active site which binds hydride, and three transport pathways for H<\/span>,2<\/sub><\/span>, H<\/span>2<\/sub><\/span>O, and electrons. H<\/span>2<\/sub><\/span> is split as part of a catalytic cycle that has been determined from synthetic model studies. [NiFe] hydrogenases could potentially be harnessed as an H<\/span>2<\/sub><\/span> synthesizer\u00a0<\/span>as an environmentally clean energy energy source. <\/span><\/p>\n

Table of Contents<\/b><\/h1>\n

Summary<\/a>
\nContext<\/a>
\nStructure and Function<\/a>
\nActive site and Catalytic Cycle<\/a>
\nProposed Canopy Model<\/a>
\nOxygen Sensitivity<\/a>
\nReferences<\/a><\/p>\n

<\/a>Context<\/b><\/h1>\n

The burning of fossil fuels releases carbon dioxide and other greenhouse gases into the atmosphere, contributing to global warming. On April 21, 2017, the atmospheric CO<\/span>2<\/sub><\/span> concentration reached 410 ppm,<\/span>\u00a0a concentration so high that the earth has not reached this level in millions of years (Figure 1)(1,2). This rise in CO<\/span>2<\/sub><\/span> concentration correlates with the rise in temperature of the earth, resulting in various consequences, for example, the melting of the ice caps. <\/span><\/p>\n

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Figure 1: Historic atmospheric CO<\/span>2<\/sub><\/span> levels.<\/span>2<\/sup><\/span><\/p>\n

Now more than ever, fossil fuels need to be abandoned and alternative energy sources need to be discovered. One promising alternative energy source is hydrogen gas. Hydrogen can be used in fuel cells to power motor vehicles. This fuel source is environmentally friendly because it yields only water as a byproduct. However, current means of producing H<\/span>2<\/span> are unsatisfactory. Steam reforming, which produces H<\/span>2<\/sub><\/span> from methane (Scheme I) also produces CO<\/span>2<\/sub><\/span> as a byproduct. <\/span><\/p>\n

Scheme I:CH<\/span>4<\/span> + H<\/span>2<\/sub><\/span>O\u21cc \u00a0CO +3H<\/span>2<\/sub><\/span><\/p>\n

CO+H<\/span>2<\/sub><\/span>O\u21cc \u00a0CO<\/span>2<\/sub><\/span>+H<\/span>2<\/sub><\/span><\/p>\n

Electrolysis of water is another alternative to producing H<\/span>2<\/span>, however this process is costly and difficult to scale up. Therefore, harnessing the power of biological catalysts to synthesize H<\/span>2<\/span> is an attractive option. [NiFe] hydrogenases are a class of enzymes that catalyze the reversible reaction shown in Scheme 2:<\/span><\/p>\n

Scheme 2: H<\/span>2<\/sub><\/span> \u21cc <\/span>\u00a0H<\/span>–<\/sup><\/span> + H<\/span>+<\/sup><\/span> \u21cc <\/span>\u00a02H<\/span>+<\/sup><\/span> + 2e<\/span>–<\/sup><\/span><\/p>\n

Therefore, the ability of [NiFe] hydrogenases to synthesize or break down H<\/span>2<\/sub><\/span> could be a useful tool for scientists developing alternative sources for H<\/span>2<\/sub><\/span>. <\/span><\/p>\n

<\/a>Structure and Function<\/b><\/h1>\n

[NiFe] hydrogenases are enzymes that catalyzes the heterolytic splitting of H<\/span>2<\/sub><\/span> (or the reverse reaction) using a catalytic center that contains one nickel and one iron atom. Because these elements are relatively accessible to humans, the Ni-Fe catalytic center is of particular interest as a potential model for a biocatalyst. <\/span><\/p>\n

[NiFe] hydrogenases are typically comprised of two subunits, a small subunit and a large subunit (some hydrogenases are membrane-bound, and contain a third subunit that is integral to the membrane). The small subunit contains three Fe-S clusters (two Fe<\/span>4<\/sub><\/span>S<\/span>4<\/sub><\/span> and one Fe3<\/sub>S<\/span>4<\/sub><\/span>) at close distances (about 10 angstroms apart). The large subunit contains the catalytic Ni-Fe center (Figure 2).<\/span><\/p>\n

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Figure 2: [NiFe] hydrogenase structure, showing large subunit in green and small subunit in blue. FeS clusters are shown in the small subunit in green and yellow (Sumner).<\/p>\n

For the enzyme to split H<\/span>2<\/span> the following must occur: H<\/span>2<\/span> diffuses in toward the buried active site; it binds to the active site and is split; protons are channeled away from the active site toward the surface of the protein; electrons are channeled away from the active site, and ultimately to a redox partner (such as a cytochrome).<\/span><\/p>\n

Three transfer pathways exist in the enzyme. The electrons flow away from the active site through the Fe-S clusters. The protons are channeled away via amino acid residue groups that can donate\/accept protons and water molecules. The H<\/span>2<\/sub><\/span> diffuses in through gas tunnels, which are lined with hydrophobic amino acids (Figure 3). These gas channels were visualized in studies with krypton, a gas ideal for investigating gas tunnels because it is hydrophobic and large. Crystallizing the [NiFe] hydrogenases together with krypton and visualizing where the krypton was bound showed that there was a hydrophobic gas tunnel through which H<\/span>2<\/sub><\/span> can diffuse to the active site.<\/span><\/p>\n

 <\/p>\n

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Figure 3: Hydrophobic gas tunnels, shown in pink, allow for gas diffusion to the active site (Kalms).<\/p>\n

<\/a>Active site and Catalytic Cycle<\/b><\/h1>\n

The Ni ion is coordinated by the thiolates of four cysteine residues (CX<\/span>2<\/sub><\/span>C on the N-terminal side of peptide, CX<\/span>2<\/sub><\/span>CX<\/span>2<\/sub><\/span>H\/R on C-terminal side), which are highly conserved. Two of the thiolates are bridging, also coordinating the Fe ion. The Fe ion is also coordinated by two CN<\/span>–<\/sup><\/span> molecules and one CO molecule. These molecules are of biogenic origin (the CN<\/span>–<\/span> comes from carbamoyl phosphate). The cyanide and carbon monoxide molecules lock the Fe in a ferrous, low-spin state (Figure 4).<\/span><\/p>\n

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Figure 4. Active site of [NiFe] hydrogenase. R509 represents an arginine residue in the second sphere that potentially plays a role in catalysis. The atom labeled “X” represents either a briding hydride (during catalysis) or a bridging oxygen atom (in enzymes inactivated by oxygen) (Brooke).<\/p>\n

The large variety of ligands (the thiolates are \u03c0-donors, the CO is a \u03c0-acceptor, and the CN<\/span>–<\/sup><\/span> is a \u03c3-donor) provides a flexible coordination environment, and allows for several stable redox states of the active site. This is important for the entry and exit of electrons at the active site.<\/span><\/p>\n

During catalysis, there is also a bridging hydride ligand that is coordinated to both the Fe and the Ni atom. This is an important part of the catalytic cycle. The catalytic cycle, while not yet fully elucidated, involves three distinct redox states called Ni-R (fully reduced), Ni-SIa<\/sub>, and Ni-C. These sites can be separated through various spectroscopy techniques.<\/span><\/p>\n

When the hydrogen molecule binds at the active site (Ni-SA<\/span>a<\/span>), it is split heterolytically, to a hydride and a proton. A basic group accepts the proton and the hydride becomes a bridging ligand to the Ni and Fe ions (forming the Ni-R state). There is debate whether the basic group is one of the cysteines in the inner sphere of coordination (this seems unlikely, however, because a thiol is not as effective a ligand). Then, the electrons are transferred from the hydride ligand to the Ni atom, and the resulting proton is released; the electrons are transferred away through the FeS clusters, reoxidizing the active site, and the original state (Ni-SI<\/span>a<\/sub><\/span>) is reformed. In the video below, the hydride ligand is shown in a model as a white sphere between the nickel and iron atoms.\u00a0<\/span><\/p>\n

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