Role of the C-terminal Gly-Gly motif of Escherichia coli MoaD, a molybdenum cofactor biosynthesis protein with a ubiquitin fold.
In Escherichia coli, the MoaD protein plays a central role in the conversion of precursor Z to molybdopterin (MPT) during molybdenum cofactor biosynthesis. MoaD has a fold similar to that of ubiquitin and contains a highly conserved C-terminal Gly-Gly motif, which in its active form contains a transferrable sulfur in the form of a thiocarboxylate group. During MPT biosynthesis, MoaD cycles between two different heterotetrameric complexes, one with MoaE to form MPT synthase and the other with MoeB, a protein similar to E1 in the ubiquitin pathway, to regenerate its transferrable sulfur. To determine the specific roles of each of the two terminal Gly residues with regard to the MoaD cycle, variants at the penultimate (Gly80) or terminal (Gly81) residues of both MoaD and thiocarboxylated MoaD were created. These variants were analyzed to determine their effects on complex formation with MoaE and MoeB, formation of the MoaD-acyl-adenylate complex, transfer of sulfur to precursor Z to form MPT, and total cofactor biosynthesis. The combined results show that while conservative substitutions at Gly80 had little effect on any of the processes that were examined, the terminal MoaD residue (Gly81) is important for transfer of sulfur to precursor Z and essential for formation of the MoaD-AMP complex. These results further our understanding of the mechanistic similarities of the MoaD-MoeB reaction to that of the ubiquitin-E1 system.
Schmitz
Jennifer
J
Institute of Biochemistry and Biology, University of Potsdam, 14476 Potsdam, Germany.
Wuebbens
Margot M
MM
Rajagopalan
K V
KV
Leimkühler
Silke
S
D020816:Amino Acid Motifs / Q000502:physiology*; D019943:Amino Acid Substitution; D002151:Calorimetry; D004926:Escherichia coli / Q000201:enzymology; D029968:Escherichia coli Proteins / Q000378:metabolism; D006033:Glycylglycine / Q000502:physiology*; D009713:Nucleotidyltransferases / Q000378:metabolism; D017510:Protein Folding; D013466:Sulfurtransferases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D025801:Ubiquitin / Q000737:chemistry*
D029968:Escherichia coli Proteins; D025801:Ubiquitin; D006033:Glycylglycine; C499585:MoeB protein, E coli; D009713:Nucleotidyltransferases; D013466:Sulfurtransferases; C085585:molybdopterin synthase