Poster abstracts
Poster number 139 submitted by Sarah Venus
Purification and characterization of cytidine-specific ribonuclease, Cusativin, for mapping nucleoside modifications in RNA
Sarah Venus (Department of Chemistry, University of Cincinnati), Patrick Limbach (Department of Chemistry, University of Cincinnati), Balasubrahmanyam Addepalli (Department of Chemistry, University of Cincinnati)
Abstract:
A cytidine-specific ribonuclease, known as Cusativin, is partially purified and characterized from cucumber (Cucumis sativus) seeds. The purification involves conventional biochemical procedures initially described by Girbés and coworkers (1). The presence of the target protein in the purified fractions was verified by its size on a denaturing polyacrylamide gel (SDS-PAGE) at different stages of purification. A 25kDa polypeptide was consistently observed on the denaturing gel following gel filtration and ion exchange chromatographic steps of purification. This partially purified protein exhibited ribonuclease activity when incubated with RNA. The digestion products were analyzed further by liquid chromatography coupled with mass spectrometry (LC-MS) to understand the specificity of RNA cleavage. The data related to nucleoside specificity and the ability of this enzyme to map nucleoside modifications in RNA will be presented. This base-specific ribonuclease can be a useful tool for mapping nucleoside modifications in RNA such as tRNA and rRNA.
References:
Rojo MA, et al. (1994) Planta 194:328-338.
Keywords: ribonuclease, cusativin, LC-MS