Evidence for a lactose-mediated association between two nuclear carbohydrate-binding proteins.
Evidence for a lactose-mediated association between two nuclear carbohydrate-binding proteins.
复制标题
两种核碳水化合物结合蛋白之间乳糖介导的关联的证据。
作者：
A. Seve;M. Felin;M. Doyennette‐Moyne;Tewfik Sahraoui;Michèle Aubery;Jean Hubert
从 HL60 细胞分离的膜耗尽的细胞核中提取 2 M NaCl 中的核蛋白。将提取的蛋白质提交至含有固定化葡萄糖、半乳糖或乳糖的亲和层析柱。通过 SDS-PAGE 解析存在于不同洗脱级分中的多肽，并进行银染色或通过分别针对葡萄糖结合蛋白 CBP67 和半乳糖结合蛋白 CBP35 和 L14 产生的单克隆或多克隆抗体进行免疫印迹分析。这里的结果显示 HL60 细胞核含有 CBP35 和 70 kDa 的葡萄糖结合凝集素 (CBP70)。这些数据说明了先前报道的含有葡萄糖基和半乳糖基残基的新糖蛋白与 HL60 细胞核的结合。此外，本研究提供了新的信息，即CBP35可以通过依赖于CBP35与乳糖的结合的相互作用与CBP70结合，并且一些亲和层析实验的结果强烈表明CBP35和CBP70通过蛋白质-蛋白质相互作用结合。这种乳糖介导的相互作用的潜在功能是根据其他人最近报道的数据进行讨论的，这些数据表明 CBP35 参与体外 mRNA 剪接，并且乳糖抑制前 RNA 底物的加工。
Nuclear proteins were extracted in 2 M NaCl from membrane-depleted nuclei isolated from HL60 cells. Extracted proteins were submitted to affinity chromatography columns containing immobilized glucose, galactose or lactose. The polypeptides present in the different eluted fractions were resolved by SDS-PAGE and were either silver stained or analysed by immunoblotting with monoclonal or polyclonal antibodies, respectively, raised against the glucose-binding protein CBP67 and the galactose-binding proteins CBP35 and L14. The results presented here show that HL60 cell nuclei contain CBP35 and a glucose-binding lectin of 70 kDa (CBP70). These data account for the previously reported binding of neoglycoproteins containing glucosyl and galactosyl residues to HL60 cell nuclei. Furthermore, the present study provides the new information that CBP35 can associate with CBP70 by interactions dependent on the binding of CBP35 to lactose, and the results of some affinity chromatography experiments strongly suggest that CBP35 and CBP70 associate by protein-protein interactions. The potential function of this lactose-mediated interaction is discussed with respect to data recently reported by others showing that CBP35 is involved in in vitro mRNA splicing and that lactose inhibits the processing of the pre-RNA substrate.