W0309

Insight Into the Role of Escherichia Coli Mobb in Molybdenum Cofactor Biosynthesis Based on the Crystal Structure. Karen McLuskey, Jennifer A. Harrison, Alexander W. Schüttelkopf, David H. Boxer and William N. Hunter, Div. of Biological Chemistry and Molecular Microbiology, School of Life Sciences, Univ. of Dundee, Dundee, UK.

Molybdoenzymes catalyze basic metabolic reactions in the carbon, nitrogen and sulfur cycles. The majority of these enzymes integrate molybdenum as the molybdenum cofactor (Moco) and in most bacterial enzymes the active form of the cofactor requires that a nucleotide moiety to be attached to Moco. Two proteins MobA and MobB, which are co-transcribed in E. coli, are involved in the attachment of the nucleotide to Moco. Although dimeric MobB is not essential to this process, it has been found to increase the activation of molybdoenzymes incorporating the cofactor by a mechanism that is not understood.

MobB displays weak GTPase activity and the primary sequence of MobB contains the Walker A motif ((G/A)-X-X-X-X-G-K-(S/T)) which is a putative nucleotide binding motif. This motif is also known as the P-loop as it is predicative of nucleotide tri-phosphate binding.

The crystal structure of MobB has been elucidated to 1.9 Å. The MobB subunit displays an unusual α/β fold arranged into a major and minor domain, the latter of which inserts between the two domains of the partner subunit creating an elongated dimer constructed around a 16 stranded β-sheet. Structural homologues of MobB were identified and found to include a number of nucleotide-binding proteins. Comparisons of these structures reveal that while the phosphate binding regions are highly conserved MobB lacks the structural elements required to interact with and efficiently bind a nucleotide base. The possibility that MobB forms a complex with the nucleotide binding MobA is also explored and modeling suggests that a MobA:MobB complex is feasible. Here we propose that MobB is an adaptor protein that acts with MobA to achieve the efficient biosynthesis of active Moco.