Manno S, Takakuwa Y, Nagao K, Mohandas N
Department of Biochemistry, Tokyo Women's Medical College, Japan.
J Biol Chem. 1995 Mar 10;270(10):5659-65. doi: 10.1074/jbc.270.10.5659.
The mechanical properties of human erythrocyte membrane are largely regulated by submembranous protein skeleton whose principal components are alpha- and beta-spectrin, actin, protein 4.1, adducin, and dematin. All of these proteins, except for actin, are phosphorylated by various kinases present in the erythrocyte. In vitro studies with purified skeletal proteins and various kinases has shown that while phosphorylation of these proteins can modify some of the binary and ternary protein interactions, it has no effect on certain other interactions between these proteins. Most importantly, at present there is no direct evidence that phosphorylation of skeletal protein(s) alters the function of the intact membrane. To explore this critical issue, we have developed experimental strategies to determine the functional consequences of phosphorylation of beta-spectrin on mechanical properties of intact erythrocyte membrane. We have been able to document that membrane mechanical stability is exquisitely regulated by phosphorylation of beta-spectrin by membrane-bound casein kinase I. Increased phosphorylation of beta-spectrin decreases membrane mechanical stability while decreased phosphorylation increases membrane mechanical stability. Our data for the first time demonstrate that phosphorylation of a skeletal protein in situ can modulate physiological function of native erythrocyte membrane.
人红细胞膜的力学性能在很大程度上受膜下蛋白质骨架的调节,其主要成分是α-和β-血影蛋白、肌动蛋白、蛋白4.1、内收蛋白和珠蛋白。除肌动蛋白外,所有这些蛋白质都可被红细胞中存在的各种激酶磷酸化。对纯化的骨架蛋白和各种激酶进行的体外研究表明,虽然这些蛋白质的磷酸化可改变一些二元和三元蛋白质相互作用,但对这些蛋白质之间的某些其他相互作用并无影响。最重要的是,目前尚无直接证据表明骨架蛋白的磷酸化会改变完整膜的功能。为了探究这一关键问题,我们开发了实验策略来确定β-血影蛋白磷酸化对完整红细胞膜力学性能的功能影响。我们已经能够证明,膜结合酪蛋白激酶I对β-血影蛋白的磷酸化可精确调节膜的力学稳定性。β-血影蛋白磷酸化增加会降低膜的力学稳定性,而磷酸化减少则会增加膜的力学稳定性。我们的数据首次证明,原位骨架蛋白的磷酸化可调节天然红细胞膜的生理功能。