Biochemical and genetic analyses of the U5, U6, and U4/U6 x U5 small nuclear ribonucleoproteins from Saccharomyces cerevisiae.
We have purified the yeast U5 and U6 pre-mRNA splicing small nuclear ribonucleoproteins (snRNPs) by affinity chromatography and analyzed the associated polypeptides by mass spectrometry. The yeast U5 snRNP is composed of the two variants of U5 snRNA, six U5-specific proteins and the 7 proteins of the canonical Sm core. The U6 snRNP is composed of the U6 snRNA, Prp24, and the 7 Sm-Like (LSM) proteins. Surprisingly, the yeast DEAD-box helicase-like protein Prp28 is stably associated with the U5 snRNP, yet is absent from the purified U4/U6 x U5 snRNP. A novel yeast U5 and four novel yeast U4/U6 x U5 snRNP polypeptides were characterized by genetic and biochemical means to demonstrate their involvement in the pre-mRNA splicing reaction. We also show that, unlike the human tri-snRNP, the yeast tri-snRNP dissociated upon addition of ATP or dATP.
Stevens
S W
SW
California Institute of Technology, Division of Biology, Pasadena 91125, USA.
Barta
I
I
Ge
H Y
HY
Moore
R E
RE
Young
M K
MK
Lee
T D
TD
Abelson
J
J
D000255:Adenosine Triphosphate / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D003080:Cold Temperature; D003838:Deoxyadenine Nucleotides / Q000378:metabolism; D005057:Eukaryotic Cells; D005656:Fungal Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism / Q000502:physiology*; D018390:Gene Targeting; D005800:Genes, Fungal; D006801:Humans; D008969:Molecular Sequence Data; D010641:Phenotype; D012322:RNA Precursors; D012326:RNA Splicing; D017414:Ribonucleoprotein, U4-U6 Small Nuclear / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D017415:Ribonucleoprotein, U5 Small Nuclear / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D017411:Ribonucleoproteins, Small Nuclear / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism / Q000502:physiology*; D012441:Saccharomyces cerevisiae / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism / Q000502:physiology*; D017386:Sequence Homology, Amino Acid; D016335:Zinc Fingers
D003838:Deoxyadenine Nucleotides; D005656:Fungal Proteins; D012322:RNA Precursors; D017414:Ribonucleoprotein, U4-U6 Small Nuclear; D017415:Ribonucleoprotein, U5 Small Nuclear; D017411:Ribonucleoproteins, Small Nuclear; D029701:Saccharomyces cerevisiae Proteins; C440569:Snu13 protein, S cerevisiae; C440570:Snu23 protein, S cerevisiae; C440571:Snu66 protein, S cerevisiae; D000255:Adenosine Triphosphate; C026600:2'-deoxyadenosine triphosphate