Rhodobacter capsulatus XdhC is involved in molybdenum cofactor binding and insertion into xanthine dehydrogenase.
Rhodobacter capsulatus xanthine dehydrogenase (XDH) is a cytoplasmic enzyme with an (alphabeta)2 heterodimeric structure that is highly identical to homodimeric eukaryotic xanthine oxidoreductases. The crystal structure revealed that the molybdenum cofactor (Moco) is deeply buried within the protein. A protein involved in Moco insertion and XDH maturation has been identified, which was designated XdhC. XdhC was shown to be essential for the production of active XDH but is not a subunit of the purified enzyme. Here we describe the purification of XdhC and the detailed characterization of its role for XDH maturation. We could show that XdhC binds Moco in stoichiometric amounts, which subsequently can be inserted into Moco-free apo-XDH. A specific interaction between XdhC and XdhB was identified. We show that XdhC is required for the stabilization of the sulfurated form of Moco present in enzymes of the xanthine oxidase family. Our findings imply that enzyme-specific proteins exist for the biogenesis of molybdoenzymes, coordinating Moco binding and insertion into their respective target proteins. So far, the requirement of such proteins for molybdoenzyme maturation has been described only for prokaryotes.
Neumann
Meina
M
Department of Proteinanalytics, Institute of Biochemistry and Biology, University of Potsdam, 14476 Potsdam, Germany.
Schulte
Marc
M
Jünemann
Nora
N
Stöcklein
Walter
W
Leimkühler
Silke
S
D002846:Chromatography, Affinity; D003067:Coenzymes / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics; D008667:Metalloproteins / Q000378:metabolism*; D000090006:Molybdenum Cofactors; D011485:Protein Binding; D011621:Pteridines / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism; D016354:Rhodobacter capsulatus / Q000201:enzymology*; D020349:Surface Plasmon Resonance; D014968:Xanthine Dehydrogenase / Q000302:isolation & purification / Q000378:metabolism*
D003067:Coenzymes; D008667:Metalloproteins; D000090006:Molybdenum Cofactors; D011621:Pteridines; D011994:Recombinant Proteins; D014968:Xanthine Dehydrogenase; C022261:molybdopterin