Role of XDHC in molybdenum cofactor insertion into xanthine dehydrogenase of Rhodobacter capsulatus

Rhodobacter capsulatus xanthine dehydrogenase (XDH) is composed of two subu nits, XDHA and XDHB, Immediately downstream of xdhB, a third gene was ident ified, designated xdhC, which is cotranscribed with xdhAB. Interposon mutag enesis revealed that the xdhC gene product is required for XDH activity. Ho wever, XDHC is not a subunit of active XDH, which forms an alpha(2)beta(2) heterotetramer in R. capsulatus. It was shown that XDHC neither is a transc riptional regulator for xdh gene expression nor influences XDH stability, T o analyze the function of XDHC for XDH in R. capsulatus, inactive XDH was p urified from an xdhC mutant strain. Analysis of the molybdenum cofactor con tent of this enzyme demonstrated that in the absence of XDHC, no molybdopte rin cofactor MPT is present in the XDHAB tetramer, In contrast, absorption spectra of inactive XDH isolated from the xdhC mutant revealed the presence of iron-sulfur clusters and flavin adenine dinucleotide, demonstrating tha t XDHC is not required for the insertion of these cofactors. The absence of MPT from XDH isolated from an xdhC mutant indicates that XDHC either acts as a specific MPT insertase or might be a specific chaperone facilitating t he insertion of MPT and/or folding of XDH during or after cofactor insertio n.