来自绿水螅多头突变体的头部激活因子受体的纯化与特性分析

Purification and characterization of the head-activator receptor from a multi-headed mutant of Chlorohydra viridissima.

作者信息

Christians S, Neubauer K H, Ulrich H

机构信息

Center for Molecular Neurobiology, Hamburg, Germany.

出版信息

FEBS Lett. 1993 Jan 25;316(2):141-6. doi: 10.1016/0014-5793(93)81203-c.

DOI:10.1016/0014-5793(93)81203-c

PMID:8380560

Abstract

In hydra the neuropeptide head activator (HA) is responsible for head-specific growth and differentiation processes. The effects of HA are mediated by high and low affinity receptors on hydra cells. In the current study HA receptors from a multi-headed mutant of Chlorohydra viridissima were solubilized from the membrane fraction using 1% Triton X-100 and 2.5 M urea. Scatchard analysis showed that the solubilized receptor had a Kd of 1.55 x 10(-9) M, indicating the low affinity subtype of the HA receptor. The solubilized receptor was purified by DMAE chromatography and subsequent affinity chromatography to homogeneity. SDS-PAGE revealed a single protein band with a molecular mass of 96 +/- 4 kDa. The native receptor eluted during gel filtration as a 113 kDa protein, and focussed with an isoelectric point of 4.8.

摘要

在水螅中，神经肽头部激活因子（HA）负责头部特异性的生长和分化过程。HA的作用是由水螅细胞上的高亲和力和低亲和力受体介导的。在当前研究中，使用1% Triton X-100和2.5 M尿素从绿水螅多头突变体的膜组分中溶解HA受体。Scatchard分析表明，溶解的受体的解离常数（Kd）为1.55×10⁻⁹ M，表明是HA受体的低亲和力亚型。通过二甲基氨基乙醇（DMAE）色谱和随后的亲和色谱将溶解的受体纯化至同质。十二烷基硫酸钠-聚丙烯酰胺凝胶电泳（SDS-PAGE）显示出一条分子量为96±4 kDa的单一蛋白带。天然受体在凝胶过滤过程中以113 kDa的蛋白形式洗脱，等电聚焦的等电点为4.8。