Purification of S1 nuclease to homogeneity and its chemical, physical and catalytic properties.

An S1 nuclease preparation was used to purify the enzyme to homogeneity. The enzyme had an isoelectric point of 4.2, and a high content of hydrophobic amino acids, especially tyrosine. It exhibited low 3'-ribonucleotidase activity. Circular dichroism analysis suggested that the contents of alpha-helix, beta-structure and random coil are 25%, 31% and 44%, respectively. The enzyme contained about 3 g atoms Zn/mol and the removal of Zn from the enzyme by addition of EDTA resulted in disruption of its secondary structure with resultant inactivation. From Con A-Sepharose chromatography, we suggest that the enzyme is a high-mannose glycoprotein. After treatment with endo-beta-N-acetylglucosaminidase H under moderate conditions, a small part of the enzyme was converted to a form lacking the sugar side chain. This form of the enzyme was as thermostable as the parent enzyme, suggesting that the sugar side chain may not be involved in thermostability of the enzyme.