Poster abstracts
Poster number 122 submitted by Chad Ratterman
Analysis of the Branchpoint Residue in Backbone Branched RNAs
Timothy Ratterman (Department of Chemistry, Carnegie Mellon University), Stephanie Mack (Department of Chemistry, Carnegie Mellon University), Subha R. Das (Department of Chemistry, Carnegie Mellon University)
Abstract:
In splicing, for intron removal, the phosphodiester of the intron 5ʹ-terminal guanosine is joined to the 2ʹ-position on an adenosine residue within the intron to form a lariat structure. Thus, excised lariat introns invariably contain a branchpoint adenosine with a guanosine as the first 2ʹ-branch residue. These lariat introns are used in myriad processes within the cell; however, first the 2ʹ-5ʹ phosphodiester bond of the lariat intron must be cleaved by lariat debranching enzyme (Dbr1p).
Preliminary data from our lab suggests that Saccharomyces cerevisiae Dbr1p can cleave backbone branched RNAs (bbRNAs) in which the branchpoint is not adenosine. Therefore, we are interested in analyzing the binding and cleavage activity of Dbr1p with such non-adenosine branchpoint containing bbRNAs. Towards this, we are synthesizing phosphoramidites for C, G and U branchpoints for bbRNAs that include dual fluorescent labels. These dual-labeled bbRNAs will enable real time kinetics assays of the Dbr1p cleavage reaction.
Keywords: Dbr1p, Backbone branched