Extracellular RNase Production and Purification from Streptomyces aureofaciens (2614)

Abstract

Streptomyces aureofaciens (NCIM 2614) culture produced extracellular RNase (291U/ml)

During screening among 9 cultures from NCIM division in MGYP medium.

 Effect of media components on RNase production showed that organism could grow in simple PG (0.8% peptone and 1% glucose) medium to replace costly MGYP medium. Medium optimization studies resulted in 2.2-fold increase in RNase activity (291-669 U/ml) and 1.6-fold increase in specific activity (5836-9211). Production profile of Streptomyces aureofaciens (2614) in PG medium (pH 7.0) at 28^οC showed that organism produces maximum RNase (669U/ml) and DNase activity (153.3U/ml) in 72 hrs and thus can be classified as non-specific nuclease. The ratio of RNase to DNase is 4: 1 suggesting that organism prefers RNA to DNA as substrate. RNase activity could be detected in the absence of metal ions while DNase activity required metal ions.

During purification of RNase enzyme, heat treatment is not a suitable method as enzyme is not thermo stable. Ammonium sulphate precipitation and dialysis resulted in 89 % recovery with 1.13-fold purification. The enzyme eluted unbound on anion exchanger (DEAE sephadex) with 46.5 % recovery which was loaded on cation exchanger (CM cellulose) and eluted with buffer containing 0.2 M salt (11% recovery). The enzyme was further loaded on gel filtration column

(FPLC Superose-12) with 11% recovery and 18.6-fold purification.