Cloning, expression and partial characterization of the C. elegans EEED8.8 gene product, a specific ADP-ribose diphosphatase, member of nudix hydrolase family

ABSTRACT

The C Elegans homologue of the human YSA1 protein, EEED and 8.8 [Nudix6], has been expressed as a thioredoxin fusion protein in Escherichia coli. It is an ADP-sugar pyrophosphatase with similar activities towards ADP-ribose and IDP-ribose. It is a specific ADP-ribose [adenosine 5-diphosphoribose] pyrophosphatase with no activities towards other nucleotides. The products of ADP-ribose hydrolysis were AMP and ribose 5-phosphate. Km and k[cat] values with ADP-ribose were 143.8 +/- 35.69 microm and18.9 +/- 2.485 micromol/min per mg protein using ADP-ribose as substrate respectively. The optimal activity was at pH 7.2 with 10 mM Mg[2+], fluoride was inhibitory, with an IC[50] of 40 microM. A major proposed function of the MutT motif proteins is to eliminate toxic nucleotide metabolites from the cell