Affinity purification of B complexes accumulating in the presence of ATPγS. (A) Splicing complexes were allowed to form on 32P-MINX pre-mRNA containing MS2 aptamers bound by MS2-MBP protein, in HeLa nuclear extract containing 2.0 mM ATPγS. Complexes were subsequently affinity purified on an amylose column and the eluate subjected to glycerol gradient centrifugation. (B) RNA composition of complexes migrating in the 60S and 40S peaks. (C) Proteins from ATPγS stalled complexes in the 40S gradient peak were separated by 2D gel electrophoresis, stained with Coomassie G-250, and the identities of protein spots were determined by mass spectrometry. (D) Summary of abundant/moderately abundant proteins present in monomeric BATPγS complexes.
