S1 nuclease of Aspergillus oryzae: characterization of the associated phosphomonoesterase activity |
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Authors: | A E Oleson E D Hoganson |
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Affiliation: | Department of Biochemistry, North Dakota State University, Fargo, North Dakota 58105 USA |
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Abstract: | S1 nuclease (EC 3.1.30.1) of Aspergillus oryzae was found to catalyze the hydrolysis of 2′- or 3′-phosphomonoester groups from several mono- and oligonucleotides. The specificity of the enzyme for mononucleotide substrates was determined by steady-state kinetic measurements at pH 4.5. The values of V were similar for all ribonucleoside 3′-phosphates tested, and they were 50–400 times greater than those for the corresponding deoxyribonucleotides or ribonucleoside 2′-phosphates. Purine nucleotides had lower apparent Km values than pyrimidine nucleotides. Apparent Km values of mononucleotides were also strongly dependent on the type of sugar and the positions of phosphoryl groups. Substrate specificity, as expressed by , occurred in the following order: ribonucleoside 3′,5′-bisphosphate > ribonucleoside 3′-phosphate > deoxyribonucleoside 3′,5'-bisphosphate > deoxyribonucleoside 3′-phosphate ≈ ribonucleoside 2′-phosphate. S1 nuclease also catalyzed the dephosphorylation of the dinucleotide ApAp at a high rate and the release of PPi from adenosine 3′-diphosphate 5′-phosphate at a low rate. The phosphomonoesterase activity of the enzyme was competitively inhibited by single-stranded DNA and 5′-nucleotides. Apparent Ki values for adenosine compounds occurred in the order ATP < ADP < AMP ? adenosine. Tests of S1 nuclease for phosphotransferase activity at pH 4.5 and 7.0 were negative. |
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Keywords: | To whom correspondence and requests for reprints should be addressed. |
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