Crystallographic and kinetic analyses of the FdsBG subcomplex of the cytosolic formate dehydrogenase FdsABG from Cupriavidus necator

Young, Tynan; Niks, Dimitri; Hakopian, Sheron; Tam, Timothy K.; Yu, Xin; Hille, Russ; Blaha, Gregor

doi:10.1074/jbc.ra120.013264

Cited by 21 publications

(36 citation statements)

References 55 publications

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“…3C, traces 1 & 2). The slow relaxation properties of this axial signal and its g-values are typical for S = ½ [2Fe-2S] 1+ clusters (37) and very similar to those detected in related FDHs from Clostridium thermoaceticum, M. trichosporum or C. necator (26,(38)(39)(40). Upon lowering the temperature to 50 K, new signals are detected peaking at g = 2.047 and 1.897 in ForCE1 (Fig.…”

Section: Epr Analysis Of Force1 and Force2supporting

confidence: 75%

Identification and characterization of a noncanonical menaquinone-linked formate dehydrogenase

Arias-Cartin

Uzel

Seduk

et al. 2022

Journal of Biological Chemistry

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The molybdenum/tungsten— bis -pyranopterin guanine dinucleotide family of formate dehydrogenases (FDHs) plays roles in several metabolic pathways ranging from carbon fixation to energy harvesting because of their reaction with a wide variety of redox partners. Indeed, this metabolic plasticity results from the diverse structures, cofactor content, and substrates used by partner subunits interacting with the catalytic hub. Here, we unveiled two noncanonical FDHs in Bacillus subtilis , which are organized into two-subunit complexes with unique features, ForCE1 and ForCE2. We show that the formate oxidoreductase catalytic subunit interacts with an unprecedented partner subunit, formate oxidoreductase essential subunit, and that its amino acid sequence within the active site deviates from the consensus residues typically associated with FDH activity, as a histidine residue is naturally substituted with a glutamine. The formate oxidoreductase essential subunit mediates the utilization of menaquinone as an electron acceptor as shown by the formate:menadione oxidoreductase activity of both enzymes, their copurification with menaquinone, and the distinctive detection of a protein-bound neutral menasemiquinone radical by multifrequency electron paramagnetic resonance (EPR) experiments on the purified enzymes. Moreover, EPR characterization of both FDHs reveals the presence of several [Fe-S] clusters with distinct relaxation properties and a weakly anisotropic Mo(V) EPR signature, consistent with the characteristic molybdenum/bis-pyranopterin guanine dinucleotide cofactor of this enzyme family. Altogether, this work enlarges our knowledge of the FDH family by identifying a noncanonical FDH, which differs in terms of architecture, amino acid conservation around the molybdenum cofactor, and reactivity.

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Section: Epr Analysis Of Force1 and Force2supporting

confidence: 75%

Identification and characterization of a noncanonical menaquinone-linked formate dehydrogenase

Arias-Cartin

Uzel

Seduk

et al. 2022

Journal of Biological Chemistry

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“…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].…”

Section: The Metal-dependent Formate Dehydrogenasesmentioning

confidence: 99%

“…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].…”

Section: The Metal-dependent Formate Dehydrogenasesmentioning

confidence: 99%

Carbon Dioxide Utilisation—The Formate Route

Maia

Moura

2020

Enzymes for Solving Humankind's Problems

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The relentless rise of atmospheric CO2 is causing large and unpredictable impacts on the Earth climate, due to the CO2 significant greenhouse effect, besides being responsible for the ocean acidification, with consequent huge impacts in our daily lives and in all forms of life. To stop spiral of destruction, we must actively reduce the CO2 emissions and develop new and more efficient “CO2 sinks”. We should be focused on the opportunities provided by exploiting this novel and huge carbon feedstock to produce de novo fuels and added-value compounds. The conversion of CO2 into formate offers key advantages for carbon recycling, and formate dehydrogenase (FDH) enzymes are at the centre of intense research, due to the “green” advantages the bioconversion can offer, namely substrate and product selectivity and specificity, in reactions run at ambient temperature and pressure and neutral pH. In this chapter, we describe the remarkable recent progress towards efficient and selective FDH-catalysed CO2 reduction to formate. We focus on the enzymes, discussing their structure and mechanism of action. Selected promising studies and successful proof of concepts of FDH-dependent CO2 reduction to formate and beyond are discussed, to highlight the power of FDHs and the challenges this CO2 bioconversion still faces.

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“…The X-ray crystal structure of the FdsBG subcomplex of the C. necator formate dehydrogenase has also been determined at a resolution of 2.3 Å [19]. This fragment of the holoenzyme contains FMN and a Fe 4 S 4 cluster in the FdsB subunit and a Fe 2 S 2 cluster in FdsG; it lacks the FdsA subunit that contains the molybdenum center and additional iron-sulfur clusters.…”

Section: The Crystal Structure Of Fdsbgmentioning

confidence: 99%

“…It is to be noted that while the N-terminal domains of FdsG and NuoE both have four helices, the first helix of FdsG runs parallel, but in NuoE, it runs across the second and third helices. The C-terminal thioredoxin-like domain of FdsG resembles the N-terminal thioredoxin-like domain of FdsB, but unlike the cofactorless FdsB domain, the FdsG domain contains a spinach ferredoxin-like Fe 2 S 2 cluster [19]. This iron-sulfur cluster is again in a hydrophobic environment near the surface, coordinated by C86 G , C91 G , C127 G , and C131 G .…”

Section: The Crystal Structure Of Fdsbgmentioning

confidence: 99%

Structure: Function Studies of the Cytosolic, Mo- and NAD+-Dependent Formate Dehydrogenase from Cupriavidus necator

et al. 2020

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Here, we report recent progress our laboratories have made in understanding the maturation and reaction mechanism of the cytosolic and NAD+-dependent formate dehydrogenase from Cupriavidus necator. Our recent work has established that the enzyme is fully capable of catalyzing the reverse of the physiological reaction, namely, the reduction of CO2 to formate using NADH as a source of reducing equivalents. The steady-state kinetic parameters in the forward and reverse directions are consistent with the expected Haldane relationship. The addition of an NADH-regenerating system consisting of glucose and glucose dehydrogenase increases the yield of formate approximately 10-fold. This work points to possible ways of optimizing the reverse of the enzyme’s physiological reaction with commercial potential as an effective means of CO2 remediation. New insight into the maturation of the enzyme comes from the recently reported structure of the FdhD sulfurase. In E. coli, FdhD transfers a catalytically essential sulfur to the maturing molybdenum cofactor prior to insertion into the apoenzyme of formate dehydrogenase FdhF, which has high sequence similarity to the molybdenum-containing domain of the C. necator FdsA. The FdhD structure suggests that the molybdenum cofactor may first be transferred from the sulfurase to the C-terminal cap domain of apo formate dehydrogenase, rather than being transferred directly to the body of the apoenzyme. Closing of the cap domain over the body of the enzymes delivers the Mo-cofactor into the active site, completing the maturation of formate dehydrogenase. The structural and kinetic characterization of the NADH reduction of the FdsBG subcomplex of the enzyme provides further insights in reversing of the formate dehydrogenase reaction. Most notably, we observe the transient formation of a neutral semiquinone FMNH·, a species that has not been observed previously with holoenzyme. After initial reduction of the FMN of FdsB by NADH to the hydroquinone (with a kred of 680 s−1 and Kd of 190 µM), one electron is rapidly transferred to the Fe2S2 cluster of FdsG, leaving FMNH·. The Fe4S4 cluster of FdsB does not become reduced in the process. These results provide insight into the function not only of the C. necator formate dehydrogenase but also of other members of the NADH dehydrogenase superfamily of enzymes to which it belongs.

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Crystallographic and kinetic analyses of the FdsBG subcomplex of the cytosolic formate dehydrogenase FdsABG from Cupriavidus necator

Cited by 21 publications

References 55 publications

Identification and characterization of a noncanonical menaquinone-linked formate dehydrogenase

Identification and characterization of a noncanonical menaquinone-linked formate dehydrogenase

Carbon Dioxide Utilisation—The Formate Route

Structure: Function Studies of the Cytosolic, Mo- and NAD+-Dependent Formate Dehydrogenase from Cupriavidus necator

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