Dumortier H, Klein Gunnewiek J, Roussel J P, van Aarssen Y, Briand J P, van Venrooij W J, Muller S
Institut de Biologie Moléculaire et Cellulaire, UPR 9021 CNRS, 15 rue Descartes, 67000 Strasbourg, France.
Nucleic Acids Res. 1998 Dec 1;26(23):5486-91. doi: 10.1093/nar/26.23.5486.
No structural information on U1C protein either in its free state or bound to the spliceosomal U1 small nuclear ribonucleoprotein (snRNP) particle is currently available. Using rabbit antibodies raised against a complete set of 15 U1C overlapping synthetic peptides (16-30 residues long) in different immunochemical tests, linear regions exposed at the surface of free and U1 snRNP-bound U1C were identified. Epitopes within at least three regions spanning residues 31-62, 85-103 and 116-159 were recognized on free and plastic-immobilized recombinant human U1C expressed in Escherichia coli, on in vitro translated U1C protein and on U1C bound to the U1 snRNP particle present in HeLa S100 extract. Using a zinc affinity labeling method, we further showed that the N-terminal U1C peptide containing a zinc-finger motif (peptide 5-34) effectively binds65Zn2+. The N-terminal region of U1C, which is functional in U1 snRNP assembly, is apparently not located at the surface of the U1 snRNP particle.
目前，关于U1C蛋白处于游离状态或与剪接体U1小核核糖核蛋白（snRNP）颗粒结合时的结构信息尚未可知。在不同的免疫化学试验中，使用针对一组完整的15个U1C重叠合成肽（长度为16 - 30个残基）产生的兔抗体，鉴定出游离的和与U1 snRNP结合的U1C表面暴露的线性区域。在大肠杆菌中表达的游离的和固定在塑料上的重组人U1C、体外翻译的U1C蛋白以及与HeLa S100提取物中存在的U1 snRNP颗粒结合的U1C上，识别出了至少三个区域（跨越第31 - 62、85 - 103和116 - 159位残基）内的表位。使用锌亲和标记方法，我们进一步表明，含有锌指基序（肽5 - 34）的U1C N端肽能有效结合65Zn2 +。在U1 snRNP组装中起作用的U1C N端区域显然不在U1 snRNP颗粒的表面。
