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The EMBO Journal (2003) 22, 1370–1380, doi:10.1093/emboj/cdg121 Figure 1 The path from nucleolar 90S to cytoplasmic 40S pre-ribosomes Thorsten Schäfer, Daniela Strau, Elisabeth Petfalski, David Tollervey and Ed Hurt

Figure 1 Enp1p is associated with 90S and 40S pre-ribosomes. (A) The sedimentation behavior of TAP-tagged Enp1p was analyzed by sucrose density gradient centrifugation. A whole-cell lysate (WCL) and fractions 5–32 from the sucrose gradient were analyzed by OD254nm measurement (upper graph) and western blotting using anti-protein A antibodies to reveal the position of TAP-tagged Enp1p (lower panel). A WCL and the pooled 40S and 90S fractions were subjected to tandem affinity purification (TAP). (B) TAP-tagged Enp1p was isolated from an unfractionated yeast WCL, from the 90S pool and the 40S pool by the TAP method. Co-purifying proteins were separated on a 4–12% SDS–polyacrylamide gradient gel, stained with Coomassie Blue and identified by mass spectrometry (see also Table I). Major non-ribosomal proteins of the 40S pre-ribosomes are marked by open circles, high molecular weight factors of the 90S pre-ribosomes by asterisks. Bands 1–29 are 90S factors identified by MS and listed in Table I. Proteins in the low molecular weight range of the gel are predominantly ribosomal Rps proteins (see also Table I).

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