Quansheng Liu, Jaclyn C. Greimann, Christopher D. Lima Cell

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Reconstitution, Activities, and Structure of the Eukaryotic RNA Exosome Quansheng Liu, Jaclyn C. Greimann, Christopher D. Lima Cell Volume 131, Issue 1, Pages 188-189 (October 2007) DOI: 10.1016/j.cell.2007.09.019 Copyright © 2007 Elsevier Inc. Terms and Conditions

Figure 1 Purification and Activities of Human Rrp41/Rrp45 and hExo9 (A) Analysis of fractions obtained after anion exchange for hExo9 (MonoQ 5/5 chromatogram; bottom panel), activity assays (top panel), and SDS-PAGE stained with Sypro-Ruby (second panel from top) or Coomassie blue (third panel from top). (B) Analysis of fractions obtained after cation exchange for hExo9 (MonoS 5/5; bottom panel), activity assays (top panel), and SDS-PAGE stained with Sypro-Ruby (second panel from top) or Coomassie blue (third panel from top). (C) Activities of wild-type and mutant hRrp41/Rrp45 and hExo9 exosomes (top panel), SDS-PAGE stained with Sypro-Ruby (middle panel) or Coomassie blue (bottom panel). Preparations in (C) were loaded at 10 μg/lane and numbered as follows: (1) wild-type hRrp41/Rrp45, (2) hRrp41(Y134R)/Rrp45, (3) hRrp41(Y134L)/Rrp45, (4) hRrp41/Rrp45(R104N,R111N), (5) hRrp41(R94V,K95A)/Rrp45, (6) hRrp41(T133A,Y134L)/Rrp45, (7) hExo9 containing wild-type hRrp41/Rrp45, (8) hExo9 containing hRrp41(T133A,Y134L)/Rrp45, (9) hExo9 containing hRrp41/Rrp45(R104N,R111N). Arrows indicate the position for Escherichia coli PNPase. Activity assays were conducted as reported (Liu et al., 2006), but with incubation for 5 min. One milligram of hExo9 was loaded onto ion exchange columns and eluted using a NaCl gradient from 50–450 mM over 25 column volumes. Cell 2007 131, 188-189DOI: (10.1016/j.cell.2007.09.019) Copyright © 2007 Elsevier Inc. Terms and Conditions