Characterization of a phosphodiesterase component in a forest soil extract

Summary

A bis-(p-nitrophenyl) phosphatase (BPN-Pase) was extracted from a forest soil and fractionated by DEAE-cellulose column chromatography into seven fractions (1, 2, 3, 4, 5, 6 and 7). The main fraction (fraction 5) was further fractionated into 3 subfraction (fractions 1, 2 and 3) by affinity chromatography for nuclease. The properties of the BPNPase in subfraction 3 were characterized and the results are reported in this article. Subfraction 3, which had a peak at about 278 run in the UV absorption spectrum, hydrolyzed 2′,3′-cyclic-nucleotides more readily than 3′,5′-cyclicnucleotides, adenylyl-(3′ → 5′)uridine, uridylyl-(3′ → 5′)adenosine, thymidine 3′-p-nitrophenyl phosphate, thymidine 5′-p-nitrophenyl phosphate, p-nitrophenyl phosphate and BPNP. Subfraction 3 hydrolyzed BPNP into 2 mol p-nitrophenyl and 1 mol inorganic phosphate during incubation. Apparent molecular weight of the BPNPase was estimated to be about 58 000 by gel filtration. The BPNPase activity had a pH optimum at 5.0 and was inhibited by Hg²⁺ and slightly inhibited by F⁻ and PO ³⁻₄ .

These observations suggest that the BPNPase is subfraction 3 has been constituted mainly with 2′,3′cyclic-nucleotide 2′-phosphodiesterase [EC 3.1.4.16] or 2′,3′-cyclic-nucleotide 3′-phosphodiesterase [EC 3.1.4.37].