CYCLIC PHOSPHODIESTERASE HAVING 3'-NUCLEOTIDASE ACTIVITY FROM BACILLUS SUBTILIS - PURIFICATION AND SOME PROPERTIES OF ENZYME

1. 1. A phosphodiesterase with 3′-nucleotidase activity was partially purified from a culture medium of Bacillus subtilis. Phosphodiesterase activity against p-dinitrophenyl phosphate was also detected in the same fraction. 2. 2. The three enzyme activities were not separated by physical and chemical procedures used so far. 3. 3. The purified enzyme preparation cleaved the phosphodiester bond of ribonucleoside 2′,3′-cyclic phosphates rapidly and that of p-dinitrophenyl phosphate slowly. It released P1 from ribonucleoside, and deoxyribonucleoside 3′-monophosphates but not from corresponding 5′- and 2′-isomers and from p-nitrophenyl phosphate. 4. 4. The final preparation was free of nonspecific acid or alkaline phosphatase, 5′-nucleotidase, phosphodiesterase against adenosine 3′,5′-cyclic monophosphate, deoxyribonuclease and ribonuclease. 5. 5. The purified enzyme was nearly homogeneous by hydroxylapatite chromatography, sedimentation, Sephadex G-200 gel filtration and polyacrylaminde gel electrophoresis. 6. 6. The s20,w was 4·93 S with an estimated molecular weight of about 50 000. 7. 7. Three enzyme activities were greatly activated by heat treatment (70°, 5 min) in the presence of Co2+ or Mn2+ (0.1 mM). This temperature-dependent activation was specific for Co2+ or Mn2+. © 1969.