利用荧光共振能量转移测量AE1阴离子交换蛋白二聚体中的距离。

Use of luminescence resonance energy transfer to measure distances in the AE1 anion exchange protein dimer.

作者信息

Knauf Philip A, Pal Prithwish

机构信息

Department of Biochemistry and Biophysics, University of Rochester Medical Center, Medical Center 2-6820, 601 Elmwood Avenue, Rochester, NY 14642, USA.

出版信息

Blood Cells Mol Dis. 2004 May-Jun;32(3):360-5. doi: 10.1016/j.bcmd.2004.01.007.

DOI:10.1016/j.bcmd.2004.01.007

PMID:15121092

Abstract

To understand how red blood cell and other proteins carry out their functions, it is necessary not only to have high-resolution crystal structures, but also to have methods that can measure changes in position of parts of the protein on the scale of Angstroms. The method of luminescence resonance energy transfer (LRET) has considerable advantages for this purpose, particularly for proteins, such as the AE1 anion exchange protein in the red cell, that are homodimers. We have applied this method, using a terbium maleimide chelate (TbM) as donor and fluorescein maleimide (FM) as acceptor, to measure the distance between the C201 residues in adjacent dimerized cytoplasmic domains of AE1 (cdAE1). The distance measured by LRET (40.8 A) corresponds closely with that calculated from the crystal structure of the cdAE1, indicating that the method can provide useful information for testing hypotheses concerning motions in this and other blood cell proteins.

摘要

为了理解红细胞和其他蛋白质如何执行其功能，不仅需要有高分辨率的晶体结构，还需要有能够在埃的尺度上测量蛋白质部分位置变化的方法。发光共振能量转移（LRET）方法在此目的上具有相当大的优势，特别是对于诸如红细胞中的AE1阴离子交换蛋白这种同型二聚体的蛋白质。我们应用了这种方法，使用铽马来酰亚胺螯合物（TbM）作为供体，荧光素马来酰亚胺（FM）作为受体，来测量AE1（cdAE1）相邻二聚化细胞质结构域中C201残基之间的距离。通过LRET测量的距离（40.8 Å）与根据cdAE1晶体结构计算出的距离密切对应，表明该方法可为测试关于这种及其他血细胞蛋白质运动的假设提供有用信息。