Cryo-EM structures reveal intricate Fe-S cluster arrangement and charging in Rhodobacter capsulatus formate dehydrogenase

Abstract

Metal-containing formate dehydrogenases (FDH) catalyse the reversible oxidation of formate to carbon dioxide at their molybdenum or tungsten active site. They display a diverse subunit and cofactor composition, but structural information on these enzymes is limited. Here we report the cryo-electron microscopic structures of the soluble Rhodobacter capsulatus FDH (RcFDH) as isolated and in the presence of reduced nicotinamide adenine dinucleotide (NADH). RcFDH assembles into a 360 kDa dimer of heterotetramers revealing a putative interconnection of electron pathway chains. In the presence of NADH, the RcFDH structure shows charging of cofactors, indicative of an increased electron load.

Carbon Dioxide Utilisation—The Formate Route

“…4, 5, 6 and 9, and references herein), show a stable hexa-coordination, with the cysteine or selenocysteine always bound to the molybdenum/tungsten ion. This is also the case of the recently solved structure of the formate-reduced D. vulgaris SeCys-W-FDH [132] and also of the NADH-reduced R. capsulatus Cys-Mo-FDH, whose structure was determined by cryo-electron microscopy [139].…”

“…6) [133,134], or the more "complex" heteromeric (abc) Mo-FDH of D. desulfuricans [135][136][137] or D. vulgaris [129,131] that contains eight redox-active centres ([4Fe-4S] centres and c-type haems) in addition to the molybdenum centre. Remarkably, the overall protein fold of the molybdenum -and tungsten-containing subunits, including the arrangement of Fe/S centre, is highly conserved 4 [116,117,119,130,132,[138][139][140].…”

“…The diversity of metal-dependent FDHs is also observed through their "molecular plasticity". Some FDHs take part in formate-hydrogen lyase systems, as is the case of FDH H from E. coli (Mo-FDH) [141], Pectobacterium atrosepticum (Mo-FDH) [142] or C. carboxidovorans (W-FDH) [143][144][145][146][147] [138] were crystallographically characterised; the R. capsulatus enzyme structure was also determined by cryo-electron microscopy [139]. In addition, also the crystallographic structure of the tungsten-containing Methanothermobacter wolfeii Nformyl-methanofuran dehydrogenase, a structurally related enzyme (see below), was solved [140].…”

“…The high K m value for formate (K m HCOO− % 55 lM; k cat HCOO − % 550 s −1 ; k HCOO− % 10 Â 10 6 M −1 s −1 ) enables D. desulfuricans SeCys-Mo-FDH to be a powerful CO 2 reducer, as long as the formate concentration is kept low (is removed from the system). In addition, once the catalysis is initiated (occurring at steady-state rates), this enzyme robustness allows the reaction to fully proceed even in the presence of dioxygen [139]. Moreover, the D. desulfuricans SeCys-Mo-FDH is also a good electrocatalyst (unmediated electrochemistry) to carry out the electrochemical reduction of CO 2 with good catalytic currents being attained [266].…”

Structural insight on the mechanism of an electron-bifurcating [FeFe] hydrogenase

“…Similar structural features have been suggested to function as a " shing rod" for catching ferredoxin in a " y casting" mechanism in cyanobacterial complex I (18) and ferredoxin NADP + reductase (19,20). Furthermore, a formate dehydrogenase enzyme that is closely related to HydABC but which does not bind ferredoxin, is speci cally missing this helix (21). Thus, we suggest that the N-terminus of HydB may be the region where ferredoxin binds and transfers electrons to the B1 cluster.…”