Schizosaccharomyces pombe Aps1, a Diadenosine 5‘,5‘ ‘‘-P¹,P⁶-Hexaphosphate Hydrolase That Is a Member of the Nudix (MutT) Family of Hydrolases:  Cloning of the Gene and Characterization of the Purified Enzyme^†,‡

The fission yeast Schizosaccharomyces pombe contains a gene on chromosome I that encodes a hypothetical nudix hydrolase, YA9E. The gene, designated aps1, has been cloned and the protein has been purified from Escherichia coli with a yield of 10 mg of Aps1/L of culture. Aps1, composed of 210 amino acids with a calculated molecular mass of 23 724 Da, behaves as a monomer with a sedimentation coefficient of 1.92 S as determined by analytical ultracentrifugation. The effective hydrodynamic radius is about 29 Å as determined by both analytical ultracentrifugation and gel-filtration chromatography. Aps1, whose expression was detected in S. pombe by Western blotting, is an enzyme that catalyzes the hydrolysis of dinucleoside oligophosphates, with Ap₆A and Ap₅A being the preferred substrates. The major reaction products are ADP and p₄A from Ap₆A and ADP and ATP from Ap₅A. Values of K_m for Ap₆A and Ap₅A are 19 μM and 22 μM, respectively, and the corresponding values of k_cat are 2.0 s^-1 and 1.7 s^-1, respectively. The enzyme has limited activity on Ap₄A and negligible activity on Ap₃A, ADP-ribose, and NADH. Aps1 catalyzes the hydrolysis of mononucleotides with decreasing activity in order from p₅A to AMP. Optimal activity with Ap₆A as substrate is observed at pH 7.6 and in the presence of 0.1−1 mM MnCl₂. Aps1 is the first nudix hydrolase isolated from S. pombe, and it is the first enzyme identified with this specific substrate specificity and reaction products.