Year of Publication
Nat Struct Mol Biol
Cryoelectron Microscopy/*methods, Hela Cells, Humans, "Imaging, Three-Dimensional", Mutation, Protein Conformation, "Ribonucleoprotein, U2 Small Nuclear/chemistry", "Ribonucleoprotein, U4-U6 Small Nuclear/chemistry", "Ribonucleoprotein, U5 Small Nuclear/c
The spliceosome is a multimegadalton RNA-protein machine that removes noncoding sequences from nascent pre-mRNAs. Recruitment of the spliceosome to splice sites and subsequent splicing require a series of dynamic interactions among the spliceosome's component U snRNPs and many additional protein factors. These dynamics present several challenges for structural analyses, including purification of stable complexes to compositional homogeneity and assessment of conformational heterogeneity. We have isolated spliceosomes arrested before the second chemical step of splicing (C complex) in which U2, U5 and U6 snRNAs are stably associated. Using electron microscopy, we obtained images of C complex spliceosomes under cryogenic conditions and determined a three-dimensional structure of a core complex to a resolution of 30 A. The structure reveals a particle of dimensions 27 x 22 x 24 nm with a relatively open arrangement of three primary domains.