Poster abstracts
Poster number 86 submitted by Stephanie Mack
Synthesis of backbone branched RNA substrates for the investigation of debranching enzyme
Stephanie Mack (Chemistry, Carnegie Mellon University), Subha R. Das (Chemistry, Carnegie Mellon University)
Abstract:
Lariat debranching enzyme (Dbr1p) cleaves the 2'-5'-phosphate linkage in spliced introns. Little is known about the exact mechanism through with Dbr1p functions and how substrate composition affects cleavage. To examine the Dbr1p mechanism, we have synthesized backbone branched RNAs (bbRNAs) that contain a 2'-5'-phoshpate linkage. Here we describe advances to the methods of bbRNA synthesis, particularly in the photodeprotection step. The stem of the bbRNA is synthesized with a 2'-photoprotected residue and the removal of this group is a critical step for branch synthesis. Using the newly optimized method, we obtained different substrates of Dbr1p that include functional group and single atom substitutions. The synthesized substrates are used for a debranching assay analysis of Dbr1p cleavage.
Keywords: Debranching Enzyme