UC Berkeley

Messenger RNA degradation is an important component of overall gene expression. During the final step of eukaryotic mRNA degradation, exoribonuclease 1 (Xrn1) carries out 5 → 3 processive, hydrolytic degradation of RNA molecules using divalent metal ion catalysis. To initiate studies of the 5 → 3 RNA decay machinery in our lab, we expressed a C-terminally truncated version of Saccharomyces cerevisiae Xrn1 and explored its enzymology using a second-generation, time-resolved fluorescence RNA degradation assay. Using this system, we quantitatively explored Xrn1s preference for 5-monophosphorylated RNA substrates, its pH dependence, and the importance of active site mutations in the molecules conserved catalytic core. Furthermore, we explore Xrn1s preference for RNAs containing a 5 single-stranded region both in an intermolecular hairpin structure and in an RNA-DNA hybrid duplex system. These results both expand and solidify our understanding of Xrn1, a centrally important enzyme whose biochemical properties have implications in numerous RNA degradation and processing pathways.