PURIFICATION AND CHARACTERIZATION OF PRE-MESSENGER-RNA SPLICING FACTOR-SF2 FROM HELA-CELLS
PURIFICATION AND CHARACTERIZATION OF PRE-MESSENGER-RNA SPLICING FACTOR-SF2 FROM HELA-CELLS
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DOI：
10.1101/gad.4.7.1158
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发表时间：
1990-07-01
影响因子：
10.5
通讯作者：
KOZAK, D
中科院分区：
文献类型：
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作者：
KRAINER, AR;CONWAY, GC;KOZAK, D
SF 2是前体mRNA体外剪接过程中5“剪接位点切割和脂质体形成所必需的活性物质，已从HeLa细胞中纯化至接近同质。最纯的组分仅含有两个33 kD的相关多肽。该部分足以补充含有剩余剪接因子的S100部分，以剪接几种前mRNA。有效剪接所需的SF 2的最佳量取决于前体mRNA底物。SF 2不同于大小相似的hnRNPA A1和U1 snRNPA多肽。内源性hnRNA与SF 2共纯化，但这种活性似乎没有必要的RNA成分。SF 2似乎是最早的特异性前螺旋体复合物的组装或稳定所必需的，尽管在缺乏其他成分的情况下，它可以以非特异性方式结合RNA。SF 2具有促进互补RNA退火的活性。因此，SF 2可以促进snRNA和前mRNA之间、互补snRNA区域之间和/或涉及分子内前mRNA螺旋的特异性RNA-RNA相互作用。其他具有RNA退火活性的纯化蛋白不能在剪接反应中取代SF 2。
SF2, an activity necessary for 5'' splice site cleavage and lariat formation during pre-mRNA splicing in vitro, has been purified to near homogeneity from HeLa cells. The purest fraction contains only two related polypeptides of 33 kD. This fraction is sufficient to complement an S100 fraction, which contains the remaining splicing factors, to splice several pre-mRNAs. The optimal amount of SF2 required for efficient splicing depends on the pre-mRNA substrate. SF2 is distinct from the hnRNP A1 and U1 snRNP A polypeptides, which are similar in size. Endogenous hnRNA copurifies with SF2, but this activity does not appear to have an essential RNA component. SF2 appears to be necessary for the assembly or stabilization of the earliest specific prespliceosome complex, although in the absence of other components, it can bind RNA in a nonspecific manner. SF2 copurifies with an activity that promotes the annealing of complementary RNAs. Thus, SF2 may promote specific RNA-RNA interactions between snRNAs and pre-mRNA, between complementary snRNA regions, and/or involving intramolecular pre-mRNA helices. Other purified proteins with RNA annealing activity cannot substitute for SF2 in the splicing reaction.