抄録
1. A method for the purification of RNase T2 [EC 2. 7. 7.17] is described, which consists of water extraction, batch-wise treatment with DEAE-cellulose, heat treatment, concentration with ammonium sulfate, DEAE-cellulose column chromatography, ethanol fractionation and DEAE-cellulose column chromatography with a milder elution gradient system.
2. By this procedure, RNase T2 was purified 900-1100-fold with a yield of about 10% (15-20 mg. with combined RNase T2-A and RNase T2-B) from 500g. of Taka-Diastase powder.
3. The purified RNase T2 was demon-strated to be homogeneous as a protein in chromatography on DEAE-cellulose, gel filtra-tion on Sephadex G-75, paper electrophoresis, sedimentation and N-terminal amino acid analysis. Furthermore, this preparation was obtained free from various contaminating enzymes in Taka-Diastase.
4. Molecular weight of RNase T2 was estimated to be 36, 000 from sedimentation equilibrium analysis and minimum molecular weight calculated from the analytical results of amino acid composition.
5. The isoelectric point was found to be around pH 5.
6. The N-terminal amino acid was determined as glutamine or glutamic acid.
7. The amino acid composition of RNase T2 was determined. This enzyme contains 6 residues of histidine, 7 residues of tryptophan and one residue of methionine.
8. Some neutral sugars were contained in the purified RNase T2.
9. Some enzymatic properties of RNase T2, such as stability, pH optimum and sensitivities toward inhibitors and activators, were investigated in detail.
10. RNase T2 was separated into RNase T2-A and RNase T2-B by the DEAE-cellulose column chromatography. RNase T2-A was indistinguishable from RNase T2-B in the enzymatic properties and the properties as a protein. The difference between the both fractions was observed only in their carbo-hydrate components.
This work has been supported by a grant to Prof. F. Egami from the Toyo Rayon Science Foundation, to which the author's thanks are due. The present author wishes to express her sincere thanks to Prof. F. Egami for his guidance and encouragement during this work.
