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Purification and characterization of an extracellular ribonuclease from a Bacillus sp. RNS3 (KX966412)

Author:
Shruti, Gupta, Sukhdev, Singh, Singh, Kanwar Shamsher
Source:
International journal of biological macromolecules 2017 v.97 pp. 440-446
ISSN:
0141-8130
Subject:
Bacillus (bacteria), RNA, ammonium sulfate, chromatography, erythrocytes, gels, hemolysis, humans, immunosuppression, inhibitory concentration 50, molecular weight, neoplasms, neurotoxicity, nucleotides, ribonucleases, viability
Abstract:
Ribonucleases (RNases) catalyze the degradation of ribonucleic acid (RNA) into smaller nucleotides. RNases display angiogenic, neurotoxic, antitumor and immunosuppressive properties. In the present study, an extracellular RNase was successfully purified to homogeneity from a Bacillus sp. RNS3 (KX966412) by salting out at 0–50% ammonium sulphate saturation followed by the gel permeation (Sephadex G-100) chromatography. The multistep purification resulted in 10.4 fold purification of RNase with a yield of 3.12%. The activity of the purified RNase was found to be 2.02U/mg protein. The purified RNase was monomeric with a molecular weight of 66kDa. It exhibited Michalis-Menten kinetics parameters Kcat 7.92min−1 and Km 0.12mg/mL. The antiproliferative activity of the purified RNase was tested against an established Hep-2C (HeLa derived) cancer cell line in vitro. The purified RNase reduced the viability of the Hep-2C cells significantly with an IC50 value of 3.53μg/mL. The haemolytic activity of purified RNase was also evaluated and unfortunately, it showed a strong haemolytic activity towards human RBCs.
Agid:
5617920