Figure 1. 3′ guanosines inhibit the Pan2 exonuclease.

a, Denaturing RNA gels showing deadenylation by recombinant S. cerevisiae Pan2–Pan3 on 5′ 6-FAM-labeled (green star) RNA substrates consisting of a 20mer non-poly(A) sequence (shown above) followed by a poly(A) tail of 30 adenosines. Where indicated, the substrate contains three additional non-A nucleotides at the 3′ end. These gels are representative of identical experiments performed 3 times. Uncropped gel images are shown in Supplementary Data Set 1. b-e, Analysis of deadenylation on poly(A) substrates with different 3′ nucleotides. Disappearance of the intact substrate was quantified by densitometry of the fluorescently labeled, full-length RNA. Data points were normalized to time = 0, and are connected by straight lines for clarity. Assays were carried out in triplicate (n = 3 independent experiments), the data points shown represent the mean, and error bars represent standard deviation. Assays are shown for full-length S. cerevisiae Pan2–Pan3 (b, e); H. sapiens PAN2–PAN3∆N278 (c); and S. cerevisiae Pan2^UCH-Exo (residues 461-1115) (d).