Isolation and characterization of the small nucleolar ribonucleoprotein particle snR30 from Saccharomyces cerevisiae.
The nucleolus of the yeast Saccharomyces cerevisiae contains the small nucleolar RNA snR30 (snoRNA), that is found associated with at least two proteins, NOP1 and GAR1. All three of these molecules are essential for the cell's viability and have been implicated in pre-rRNA maturation. NOP1 and GAR1 are believed to be general rRNA-processing factors or, alternatively, integral protein components of the small nucleolar ribonucleoprotein particle snR30 (snoRNP). In this paper, we describe procedures for the biochemical isolation of snR30 RNP, and we identify seven snR30 RNP proteins of molecular masses of 10, 23, 25, 38, 46, 48, and 65 kDa, including the previously reported GAR1 protein. Additional proteins, including NOP1, may also be components of snR30 RNP but are lost during our stringent isolation procedure. The 10-, 23-, and 25-kDa (GAR1) and 65-kDa proteins remain tightly associated with the snR30 RNA even after isopycnic sedimentation in cesium sulfate gradients. Electron microscopy of Mono Q-purified snR30 RNPs show a slightly elongated two-domain structure approximately 20 nm long and 14 nm wide.
Lübben
B
B
Institut für Molekularbiologie und Tumorforschung, Philipps Universität Marburg, Federal Republic of Germany.
Fabrizio
P
P
Kastner
B
B
Lührmann
R
R
D001483:Base Sequence; D015153:Blotting, Western; D002466:Cell Nucleolus / Q000378:metabolism; D002846:Chromatography, Affinity; D002852:Chromatography, Ion Exchange; D003001:Cloning, Molecular; D004591:Electrophoresis, Polyacrylamide Gel; D008854:Microscopy, Electron; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009838:Oligodeoxyribonucleotides; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000302:isolation & purification / Q000648:ultrastructure; D017411:Ribonucleoproteins, Small Nuclear / Q000302:isolation & purification* / Q000648:ultrastructure; D012441:Saccharomyces cerevisiae / Q000378:metabolism*
D009838:Oligodeoxyribonucleotides; D011994:Recombinant Proteins; D017411:Ribonucleoproteins, Small Nuclear