The chaperone FdsC for Rhodobacter capsulatus formate dehydrogenase binds the bis‐molybdopterin guanine dinucleotide cofactor

Abstract

Edited by Miguel De la RosaKeywords: Molybdenum cofactor L-cysteine desulfurase Formate dehydrogenase Chaperone bis-MGD a b s t r a c t Molybdoenzymes are complex enzymes in which the molybdenum cofactor (Moco) is deeply buried in the enzyme. Most molybdoenzymes contain a specific chaperone for the insertion of Moco. For the formate dehydrogenase FdsGBA from Rhodobacter capsulatus the two chaperones FdsC and FdsD were identified to be essential for enzyme activity, but are not a subunit of the mature enzyme. Here, we purified and characterized the FdsC protein after heterologous expression in Escherichia coli. We were able to copurify FdsC with the bound Moco derivate bis-molybdopterin guanine dinucleotide. This cofactor successfully was used as a source to reconstitute the activity of molybdoenzymes. Structured summary of protein interactions:FdsC and FdsC bind by molecular sieving (View interaction) FdsD binds to RcMobA by surface plasmon resonance (View interaction) FdsC binds to RcMobA by surface plasmon resonance (View interaction) FdsC binds to FdsA by surface plasmon resonance (View interaction)

Sulphur shuttling across a chaperone during molybdenum cofactor maturation

“…As nucleotides are chemical moieties of the Mo-bisPGD structure 14 , it has been hypothesized that EcFdhD could bind and sulphurate the cofactor before its insertion into FDHs 11 . A recent study supports this hypothesis with the copurification of Mo-bisPGD onto FdsC, an FdhD homologue from Rhodobacter capsulatus 15 .…”

“…It is noteworthy that this archaeon appears to lack a cysteine desulphurases 18 , which raise the possibility that the FdhD homologue functions as a chaperone only and does not participate in the sulphuration of Mo-bisPGD. In the case of the R. capsulatus FdsC, the sequence harbours two cysteine residues within the loop, but their substitutions for alanine residues did not impair R. capsulatus FDH activity in E. coli 15 . FdsC was shown to bind Mo-bisPGD, consistent with conservation of the residues involved in Mo-bisPGD binding identified in our study.…”

The role of FeS clusters for molybdenum cofactor biosynthesis and molybdoenzymes in bacteria

“…The terminal electron acceptor for formate oxidation is specifically NAD + (NADP + is not used), which accepts the two electrons from the FMN cofactor. Since in the absence of the chaperones FdsC and FdsD during expression, a heterotrimeric enzyme lacking Moco but containing the set of [FeS] clusters and FMN is formed, it is believed that bis-MGD is inserted into the enzyme after the insertion of the other cofactors into the heterotrimer [113]. So far, the FeS clusters have not been further characterized in R. capsulatus FDH and nothing is known about the proteins that specifically insert the FeS clusters or FMN into FdsGBA.…”

The Biosynthesis of the Molybdenum Cofactor inEscherichia coliand Its Connection to FeS Cluster Assembly and the Thiolation of tRNA

“…Recently, it was shown that these chaperones not only facilitate the insertion of bis-MGD into the target enzyme, but also directly bind bis-MGD [88]. This was shown by the FdsC-FdsA system for maturation of R. capsulatus formate dehydrogenase (FDH).…”

“…While FdsD has only counterparts in some oxygen-tolerant FDHs, FdsC shares high amino acid sequence homologies to E. coli FdhD, the chaperone for the membrane-bound FDH (FdhF) [84]. FdsC was directly copurified with bound bis-MGD [88]. The definitive proof that bis-MGD was bound to FsdC and not only MGD was given by the reconstitution of E. coli TMAO reductase activity in a system solely consisting of FdsC and apo-TMAO reductase.…”