The binding of nucleotides to 3'-nucleotidase from wheat germ

Abstract

The 3'-mononucleotidase (3'-ribonucleotide phosphohydrolase, EC 3.1.3.6) from wheat germ has been purified 2,000 fold. The enzyme has a molecular weight of approximatley 32,000, as judged by the use of G-100 gel filtration, and does not attack 2'- or 5'- nucleotides. In order to obtain some indications on the structural requirements for binding and reactivity, the purified enzyme has been subjected to kinetic analyses, including initial velocities with several 3'-ribomononucleotides, inhibition by 5'- nucleotides and nucleotide-analogues, and effect of pH and sulphydryl compounds. The data indicate one base binding site at the active site of the enzyme. This site appears to be the same involved in the binding of both substrates and inhibitors, with higher affinity for purine nucleotides than for pyrimidine compounds, in the order guanosine greater than adenosine greater than inosine greater than uridine greater than cytidine nucleotides