Ribonuclease A (RNase A) from Bovine Pancreas

• Ribonuclease A (RNase A) belongs to an endoribonuclease class of ribonucleases. In contrast to exoribonucleases that cleave/degrade RNA in 3’-5’ direction, endoribonucleases degrade RNA endoribonucleolytically in 5’-3’ direction.
• RNase A is a digestive enzyme which is secreted by the pancreas to digest RNA. It is abundantly present in the pancreas, therefore, the pancreas is a valuable source for RNase A.
• Mature bovine pancreatic RNase A only has 124 amino acids with a molecular weight of 13.7 kDa. It lacks tryptophan amino acid.
• In contrast to other known members of endoribonuclease, RNase A is not a glycoprotein.
• RNase A is active under a wide range of reaction conditions (temperature range 15 – 70°C; pH range 6–10). The optimal temperature for its activity is 60°C and the optimal pH is 7.6.
• RNase A is quite stable to both heat and detergents.
• It cleaves both single-stranded and double-stranded RNA as well the RNA strand in RNA-DNA hybrids at a low salt concentration (0 to 100 mM NaCl). However, it specifically cleaves single-stranded RNA at the higher salt concentration (0.3M NaCl or higher)

RNase A stock solution:

• Generally, a 10 mg/ml RNase A solution is prepared in 10 mM Tris.Cl (pH 7.5) or TE [(10 mM Tris.Cl (pH 7.6), 1 mM EDTA]. The stock is stable for at least 1 year at -20°C.
• Most suppliers provide molecular biology grade RNase A powder which is free from any DNase activity. Sometimes, a low level of DNase activity in RNase stock solution can be detected which can be easily eliminated by incubating the stock solution in boiling water bath for 5 to 10 min (see protocol). Boiling RNase A solution for a short time does not inactivate RNase A but is sufficient to inactivate DNase activity.
• If the RNase A stock is suspected to have high DNase activity, it is recommended to prepare a 10 mg/ml stock solution in sodium acetate (pH 5.2), incubate the solution in a boiling water bath for 20 – 30 min, then adjust the pH with 1M Tris.Cl (pH 7.5). RNase A is comparatively very stable at low pH (between pH 2.0 – 4.5).