Substrate Specificity of Ribonuclease from Aspergillus saitoi

Abstract

1. The substrate specificity of Ribonuclease M [EC 2.7.7.17, Ribo-nucleate nucleotido-2'-transferase (cyclizing)], which was obtained from Aspergillus saitoi, was studied in detail using several kinds of RNA's and sixteen diribonucleoside monophosphates (XpY's, where X or Y indicates one of the four main ribonucleoside residues), as the substrates. 2. The K_m value of this enzyme for the XpY's differed depending on the nucleoside residues, X or Y. With different nucleoside residues the order of the K_m values was: uridine>guanosine>cytidine>adenosine. On the other hand, the maximum velocity (V_max) for XpY was lowest when either of X or Y was adenosine. 3. It was found that, when mixtures of several XpY's were used as substrates of this enzyme, the XpY's giving the smallest K_m value with the enzyme were split firstest. 4. A spectrophotometric procedure was developed to follow the enzymatic transesterification of the substrates, XpY's, with purine nucleo-side as X. 5. Similar kinetic studies were made on ribonuclease T₂ [EC 2.7.7.17] from Aspergillus orizae.