The Carbon Monoxide Dehydrogenase from Desulfovibrio vulgaris

Abstract

Ni-containing Carbon Monoxide Dehydrogenases (CODHs) catalyze the reversible conversion between CO and CO₂and are involved in energy conservation and carbon fixation. These homodimeric enzymes house two NiFeS active sites (C-clusters) and three accessory [4Fe-4S] clusters. The Desulfovibrio vulgaris (Dv) genome contains a two-gene CODH operon coding for a CODH (cooS) and a maturation protein (cooC) involved in nickel insertion in the active site. According to the literature, the question of the precise function of CooC as a chaperone folding the C-cluster in a form which accommodates free nickel or as a mere nickel donor is not resolved. Here, we report the biochemical and spectroscopic characterization of two recombinant forms of the CODH, produced in the absence and in the presence of CooC, designated CooS and CooS(C), respectively. CooS contains no nickel and cannot be activated, supporting the idea that the role of CooC is to fold the C-cluster so that it can bind nickel. As expected, CooS(C) is Ni-loaded, reversibly converts CO and CO₂, displays the typical Cred1 and Cred2 EPR signatures of the C-cluster and activates in the presence of methyl viologen and CO in an autocatalytic process. However, Ni-loaded CooS(C) reaches maximum activity only upon reductive treatment in the presence of exogenous nickel, a phenomenon that had not been observed before. Surprisingly, the enzyme displays the Cred1 and Cred2 signatures whether it has been activated or not, showing that this activation process of the Ni-loaded Dv CODH is not associated with structural changes at the active site.

Heterologous Expression of the Clostridium carboxidivorans CO Dehydrogenase Alone or Together with the Acetyl Coenzyme A Synthase Enables both Reduction of CO 2 and Oxidation of CO by Clostridium acetobutylicum

“…The CODH/ACS complex has been studied in several acetogens, such as M. thermoacetica, C. formicoaceticum, and Acetobacterium woodii, as well as Desulfovibrio vulgaris and Carboxydothermus hydrogenoformans (9,28,36,41,42). In these organisms, a reduced anaerobic environment provides the conditions for CO 2 fixation (43).…”

“…The CODH/ACS protein complex in acetogens couples the reduction of CO 2 to CO with the subsequent condensation with a methyl group to form acetyl-CoA (28). The CO which is generated at the C-cluster of the CODH travels through the internal channel to the ACS subunit and catalytic active site, relying on a tight association between the two subunits.…”

Reliable estimation of the kinetic parameters of redox enzymes by taking into account mass transport towards rotating electrodes in protein film voltammetry experiments

“…Distinct CODHs have been studied using PFV in the past decade, such as CODH I and II from Carboxydothermus hydrogenoformans or CODH from Desulfovibrio vulgaris [17][18][19][20]. However, the high activity of CODHs introduces a complication for their study using PFV, since many of the electrochemical signals we recorded with Desulfovibrio vulgaris CODH adsorbed onto pyrolytic graphite edge electrodes showed signs of limitation by transport, even when rotating the electrode at 6 krpm; we thus decided to include transport limitation in our modelling.…”

AcsF Catalyzes the ATP-dependent Insertion of Nickel into the Ni,Ni-[4Fe4S] Cluster of Acetyl-CoA Synthase

“…Additional proteins support the maturation of the CODH in Rhodospirillum rubrum (25); however, these additional proteins are not conserved in bacteria outside the Rhodospirillaceae and may be functionally replaced by non-homologous isofunctional enzymes in other organisms. Although CooC was shown to mature monofunctional CODHs (25)(26)(27)(28), it was proposed that the homologous ATPase AcsF supports the assembly of the C-cluster in the bifunctional ACS-CODH complex in Moorella thermoacetica (33).…”

“…CODH activity in vivo depends on an ATPase termed CooC (25)(26)(27)(28). CooC belongs to the MinD-type ATPases, an enzyme family with diverse functions and a synapomorphic KGG signature in their Walker A motif (GKGGhGK(T/S)) containing a characteristic lysine residue (signature lysine) in addition to the P-loop lysine (29).…”