Thermophilic Polynucleotide Phosphorylase from Thermus thermophilus

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Abstract
A thermophilic polynucleotide phosphorylase lacking polynucleotide phosphorolytic activity was purified from Thermus thermophilus HB‐8 strain. The enzyme is an altered form of the native polynucleotide phosphorylase, probably attacked by the proteinase(s) of this extreme thermophile during the purification process. This modified enzyme lacks phosphorolytic activity to poly(A) while retaining weak activity to phosphorolyse tetranucleotides or hexanucleotides. The purified enzyme was shown to be homogeneous by electrophoretic analysis in polyacrylamide gel. This enzyme had a molecular weight of 190000 as calculated both from electrophoresis on polyacrylamide gel and from the Stokes' radius derived from the gel filtration pattern and the sedimentation coefficient. The enzyme was separated into three polypeptide chains by polyacrylamide gel electrophoresis in the presence of sodium dodecylsulphate; their molecular weights were calculated to be 92000, 73000 and 35000. The enzyme was thermophilic and thermotolerant, exhibiting its maximal activity at 70 °C. The four ribonucleoside diphosphates (ADP, GDP, UDP and CDP) were polymerized to the extent of 7‐S size.

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