Purification, characterization and cytotoxic properties of a bacterial RNase.

Rakesh Kumar, Sunita Katwal, Bhupender Sharma, Abhishek Sharma, Sanjeev Puri, Nidhi Kamboj, Shamsher Singh Kanwar

International journal of biological macromolecules 2021 Jan 01

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An RNase produced by Bacillus safensis RB-5 was purified up to 22.32-fold by successive techniques of salting out, DEAE-anion exchange and gel permeation (Sephadex G-100) chromatography techniques with a yield of 2.27%. The purified RNase possessed a single band in SDS-PAGE (Mr ~ 60 kDa). The purified RNase showed optimal activity at temperature of 37 °C and pH 7.5 in the presence of substrate (Yeast RNA) and Mg2+ ions. The RNase activity was strongly inhibited by Hg2+ and mildly by Fe2+, Ba2+ and Zn2+ ions. Its half-life was found to be 8 h at 37 °C. The RNase kinetics study showed Km and Vmax value of 0.3 mM and 9.2 μmol/mg/min, respectively. The purified RNase also showed cytotoxic and antiproliferative activities towards a few transformed cell lines. The purified RNase (IC50 0.035 U/mL) effectively inhibited RD and Hep-2C cells proliferation & migration, while sparing HEK 293 cells. The purified RNase was cytotoxic as well as effective degrader of the RNA of transformed RD cells at low concentration. Moreover, the purified RNase of B. safensis RB-5 was found to possess a little hemolytic activity towards human RBCs. Copyright © 2020 Elsevier B.V. All rights reserved.

Citation

Rakesh Kumar, Sunita Katwal, Bhupender Sharma, Abhishek Sharma, Sanjeev Puri, Nidhi Kamboj, Shamsher Singh Kanwar. Purification, characterization and cytotoxic properties of a bacterial RNase. International journal of biological macromolecules. 2021 Jan 01;166:665-676