Redox-dependent rearrangements of the NiFeS cluster of carbon monoxide dehydrogenase (original) (raw)
Related papers
Crystal Structure of a Carbon Monoxide Dehydrogenase Reveals a [Ni-4Fe-5S] Cluster
Science, 2001
The homodimeric nickel-containing CO dehydrogenase from the anaerobic bacterium Carboxydothermus hydrogenoformans catalyzes the oxidation of CO to CO 2 . A crystal structure of the reduced enzyme has been solved at 1.6 angstrom resolution. This structure represents the prototype for Ni-containing CO dehydrogenases from anaerobic bacteria and archaea. It contains five metal clusters of which clusters B, BЈ, and a subunit-bridging, surface-exposed cluster D are cubane-type [4Fe-4S] clusters. The active-site clusters C and CЈ are novel, asymmetric [Ni-4Fe-5S] clusters. Their integral Ni ion, which is the likely site of CO oxidation, is coordinated by four sulfur ligands with square planar geometry.
Carbon Monoxide Induced Decomposition of the Active Site [Ni−4Fe−5S] Cluster of CO Dehydrogenase
Journal of the American Chemical Society, 2004
During the past two years, crystal structures of Cu-and Mo-containing carbon monoxide dehydrogenases (CODHs) and Ni-and Fe-containing CODHs have been reported. The active site of CODHs from anaerobic bacteria (cluster C) is composed of Ni, Fe, and S for which crystallographic studies of the enzymes from Carboxydothermus hydrogenoformans, Rhodospirillum rubrum, and Moorella thermoacetica revealed structural similarities in the overall protein fold but showed substantial differences in the essential Ni coordination environment. The [Ni-4Fe-5S] cluster C in the fully catalytically competent dithionitereduced CODH II from C. hydrogenoformans (CODHIICh) at 1.6 Å resolution contains a characteristic µ2sulfido ligand between Ni and Fe1, resulting in a square-planar ligand arrangement with four S-ligands at the Ni ion. In contrast, the [Ni-4Fe-4S] clusters C in CO-treated CODH from R. rubrum resolved at 2.8 Å and in CO-treated acetyl-CoA synthase/CODH complex from M. thermoacetica at 2.2 and 1.9 Å resolution, respectively, do not contain the µ2-sulfido ligand between Ni and Fe1 and display dissimilar geometries at the Ni ion. The [Ni-4Fe-4S] cluster is composed of a cubane [Ni-3Fe-4S] cluster linked to a mononuclear Fe site. The described coordination geometries of the Ni ion in the [Ni-4Fe-4S] cluster of R. rubrum and M. thermoacetica deviate from the square-planar ligand geometry in the [Ni-4Fe-5S] cluster C of CODHIICh. In addition, the latter was converted into a [Ni-4Fe-4S] cluster under specific conditions. The objective of this study was to elucidate the relationship between the structure of cluster C in CODHIICh and the functionality of the protein. We have determined the CO oxidation activity of CODHIICh under different conditions of crystallization, prepared crystals of the enzyme in the presence of dithiothreitol or dithionite as reducing agents under an atmosphere of N2 or CO, and solved the corresponding structures at 1.1 to 1.6 Å resolutions. Fully active CODHIICh obtained after incubation of the enzyme with dithionite under N2 revealed the [Ni-4Fe-5S] cluster. Short treatment of the enzyme with CO in the presence of dithiothreitol resulted in a catalytically competent CODHIICh with a CO-reduced [Ni-4Fe-5S] cluster, but a prolonged treatment with CO caused the loss of CO-oxidizing activity and revealed a [Ni-4Fe-4S] cluster, which did not contain a µ2-S. These data suggest that the [Ni-4Fe-4S] cluster of CODHIICh is an inactivated decomposition product originating from the [Ni-4Fe-5S] cluster.
ACS Catalysis, 2020
Ni−Fe CO-dehydrogenases (CODHs) catalyze the conversion between CO and CO 2 using a chain of Fe−S clusters to mediate long-range electron transfer. One of these clusters, the Dcluster, is surface-exposed and serves to transfer electrons between CODH and external redox partners. These enzymes tend to be extremely O 2-sensitive and are always manipulated under strictly anaerobic conditions. However, the CODH from Desulfovibrio vulgaris (Dv) appears unique: exposure to micromolar concentrations of O 2 on the minutes-time scale only reversibly inhibits the enzyme, and full activity is recovered after reduction. Here, we examine whether this unusual property of Dv CODH results from the nature of its D-cluster, which is a [2Fe-2S] cluster, instead of the [4Fe-4S] cluster observed in all other characterized CODHs. To this aim, we produced and characterized a Dv CODH variant where the [2Fe-2S] D-cluster is replaced with a [4Fe-4S] D-cluster through mutagenesis of the D-cluster−binding sequence motif. We determined the crystal structure of this CODH variant to 1.83-Å resolution and confirmed the incorporation of a [4Fe-4S] D-cluster. We show that upon long-term O 2-exposure, the [4Fe-4S] Dcluster degrades, whereas the [2Fe-2S] D-cluster remains intact. Crystal structures of the Dv CODH variant exposed to O 2 for increasing periods of time provide snapshots of [4Fe-4S] D-cluster degradation. We further show that the WT enzyme purified under aerobic conditions retains 30% activity relative to a fully anaerobic purification, compared to 10% for the variant, and the WT enzyme loses activity more slowly than the variant upon prolonged aerobic storage. The D-cluster is therefore a key site of irreversible oxidative damage in Dv CODH, and the presence of a [2Fe-2S] D-cluster contributes to the O 2-tolerance of this enzyme. Together, these results relate O 2-sensitivity with the details of the protein structure in this family of enzymes.
In view of the depletion of fossil fuel reserves and climatic effects of greenhouse gas emissions, Ni,Fe-containing carbon monoxide dehydrogenase (Ni-CODH) enzymes have attracted increasing interest in recent years for their capability to selectively catalyze the reversible reduction of CO 2 to CO (CO 2 + 2H + + 2e − F CO + H 2 O). The possibility of converting the greenhouse gas CO 2 into useful materials that can be used as synthetic building blocks or, remarkably, as carbon fuels makes Ni-CODH a very promising target for reverse-engineering studies. In this context, in order to provide insights into the chemical principles underlying the biological catalysis of CO 2 activation and reduction, quantum mechanics calculations have been carried out in the framework of density functional theory (DFT) on different-sized models of the Ni-CODH active site. With the aim of uncovering which stereoelectronic properties of the active site (known as the C-cluster) are crucial for the efficient binding and release of CO 2 , different coordination modes of CO 2 to different forms and redox states of the C-cluster have been investigated. The results obtained from this study highlight the key role of the protein environment in tuning the reactivity and the geometry of the Ccluster. In particular, the protonation state of His93 is found to be crucial for promoting the binding or the dissociation of CO 2. The oxidation state of the C-cluster is also shown to be critical. CO 2 binds to C red2 according to a dissociative mechanism (i.e., CO 2 binds to the C-cluster after the release of possible ligands from Fe u) when His93 is doubly protonated. CO 2 can also bind noncatalytically to C red1 according to an associative mechanism (i.e., CO 2 binding is preceded by the binding of H 2 O to Fe u). Conversely, CO 2 dissociates when His93 is singly protonated and the C-cluster is oxidized at least to the C int redox state.
Proceedings of the National Academy of Sciences, 2001
A crystal structure of the anaerobic Ni-Fe-S carbon monoxide dehydrogenase (CODH) from Rhodospirillum rubrum has been determined to 2.8-Å resolution. The CODH family, for which the R. rubrum enzyme is the prototype, catalyzes the biological oxidation of CO at an unusual Ni-Fe-S cluster called the C-cluster. The Ni-Fe-S C-cluster contains a mononuclear site and a four-metal cubane. Surprisingly, anomalous dispersion data suggest that the mononuclear site contains Fe and not Ni, and the four-metal cubane has the form [NiFe 3 S 4 ] and not [Fe 4 S 4 ]. The mononuclear site and the four-metal cluster are bridged by means of Cys 531 and one of the sulfides of the cube. CODH is organized as a dimer with a previously unidentified [Fe 4 S 4 ] cluster bridging the two subunits. Each monomer is comprised of three domains: a helical domain at the N terminus, an α/β (Rossmann-like) domain in the middle, and an α/β (Rossmann-like) domain at the C terminus. The helical domain contributes ligands ...
Biochemistry, 2009
Nickel-containing carbon monoxide dehydrogenases (CODHs) reversibly catalyze the oxidation of carbon monoxide to carbon dioxide and are of vital importance in the global carbon cycle. The unusual catalytic CODH C-cluster has been crystallographically characterized as either a NiFe 4 S 4 or a NiFe 4 S 5 metal center, the latter containing a fifth, additional sulfide that bridges Ni and a unique Fe site. To determine whether this bridging sulfide is catalytically relevant and to further explore the mechanism of the C-cluster, we obtained crystal structures of the 310 kDa bifunctional CODH/acetyl-CoA synthase complex from Moorella thermoacetica bound both with a substrate H 2 O/OHmolecule and with a cyanide inhibitor. X-ray diffraction data were collected from native crystals and from identical crystals soaked in a solution containing potassium cyanide. In both structures, the substrate H 2 O/OHmolecule exhibits binding to the unique Fe site of the C-cluster. We also observe cyanide binding in a bent conformation to Ni of the C-cluster, adjacent the substrate H 2 O/OHmolecule. Importantly, the bridging sulfide is not present in either structure. As these forms of the C-cluster represent the coordination environment immediately before the reaction takes place, our findings do not support a fifth, bridging sulfide playing a catalytic role in the enzyme mechanism. The crystal structures presented here, along with recent structures of CODHs from other organisms, have led us toward a unified mechanism for CO oxidation by the C-cluster, the catalytic center of an environmentally important enzyme. † This work is supported by NIH Grants GM69857 (C.L.D.) and GM39451 (S.W.R.) and the MIT Energy Initiative. ‡ The atomic coordinates and structure factors (PDB accession numbers 3I01 and 3I04) have been deposited in the Protein Data Bank,
Proceedings of the National Academy of Sciences of the United States of America, 2002
The CO dehydrogenase of the eubacterium Oligotropha carboxidovorans is a 277-kDa Mo-and Cu-containing iron-sulfur flavoprotein. Here, the enzyme's active site in the oxidized or reduced state, after inactivation with potassium cyanide or with n-butylisocyanide bound to the active site, has been reinvestigated by multiple wavelength anomalous dispersion measurements at atomic resolution, electron spin resonance spectroscopy, and chemical analyses. We present evidence for a dinuclear heterometal [CuSMo(AO)OH] cluster in the active site of the oxidized or reduced enzyme, which is prone to cyanolysis. The cluster is coordinated through interactions of the Mo with the dithiolate pyran ring of molybdopterin cytosine dinucleotide and of the Cu with the S␥ of Cys-388, which is part of the active-site loop VAYRC 388 SFR. The previously reported active-site structure [Dobbek, H., Gremer, L., Meyer, O. & Huber, R. (1999) Proc. Natl.
Proceedings of the …, 1992
The nickel/iron/sulfur center of the carbon monoxide dehydrogenase (carbon monoxide:(acceptor)oxidoreductase; EC 1.2.99.2) enzyme from Rhodospirillum rubrum (Rr-CODH) was studied by x-ray absorption spectroscopy at the Ni K edge. Extended x-ray absorption fine structure data show that the first Ni coordination shell consists of 2 S atoms at 2.23 A and 2-3 N/O atoms at 1.87 A. The edge structure indicates a distorted tetrahedral or five-coordinate Ni environment in both oxidized and reduced Rr-CODH. By comparing second-shell extended x-ray absorption fine structure data of Rr-CODH to that of (Et4N)3[NiFe3S4(SEt)4], a cubane-type cluster, it was clearly established that Ni in the Rr-CODH center is not involved in the core of a NiFe3S4 cubane cluster. One model consistent with the results is a mononuclear
JBIC Journal of Biological Inorganic Chemistry, 2005
Carbon monoxide dehydrogenase (CODH) from Rhodospirillum rubrum catalyzes the oxidation of CO to CO 2 . A unique [NiFe 4 S 4 ] cluster, known as the Ccluster, constitutes the active site of the enzyme. When grown in Ni-deficient medium R. rubrum accumulates a Ni-deficient apo form of CODH that is readily activated by Ni. It has been previously shown that activation of apo-CODH by Ni is a two-step process involving the rapid formation of an inactive apo-CODH•Ni complex prior to conversion to the active holo-CODH. We have generated CODH variants with substitutions in cysteine residues involved in the coordination of the [Fe 3 S 4 ] portion of the C-cluster. Analysis of the variants suggests that the cysteine residues at positions 338, 451, and 481 are important for CO oxidation activity catalyzed by CODH but not for Ni binding to the C-cluster. C451S CODH is the only new variant that retains residual CO oxidation activity. Comparison of the kinetics and pH dependence of Ni activation of the apo forms of wildtype, C451S, and C531A CODH allowed us to develop a model for Ni insertion into the C-cluster of CODH in which Ni reversibly binds to the C-cluster and subsequently coordinates Cys531 in the rate-determining step.