通过免疫沉淀法研究Bak/Bax激活构象变化。

Investigating Bak/Bax activating conformation change by immunoprecipitation.

作者信息

Dewson Grant

机构信息

Cell Signalling and Cell Death Division, Walter and Eliza Hall Institute of Medical Research, Parkville, Melbourne, Victoria 3052, Australia;Department of Medical Biology, The University of Melbourne, Melbourne, Victoria 3010, Australia.

出版信息

Cold Spring Harb Protoc. 2015 May 1;2015(5):472-6. doi: 10.1101/pdb.prot086454.

DOI:10.1101/pdb.prot086454

PMID:25934937

Abstract

Activation of both Bax and Bak during apoptosis involves significant conformation change. Investigation of this phenomenon by immunoprecipitation (IP) requires a detergent such as CHAPS that does not induce significant conformation change. IP with conformation-specific Bax or Bak antibodies is observed in CHAPS only following an apoptotic stimulus, whereas the same antibodies will immunoprecipitate from both nonapoptotic and apoptotic cells in the presence of Triton X-100. Thus, the latter detergent can serve as a positive control for IP, as described here.

摘要

细胞凋亡过程中 Bax 和 Bak 的激活涉及显著的构象变化。通过免疫沉淀（IP）研究这一现象需要一种不会诱导显著构象变化的去污剂，如 CHAPS。仅在凋亡刺激后，在 CHAPS 中才能观察到用构象特异性 Bax 或 Bak 抗体进行的免疫沉淀，而在 Triton X-100 存在的情况下，相同的抗体能从非凋亡细胞和凋亡细胞中都进行免疫沉淀。因此，如本文所述，后一种去污剂可作为 IP 的阳性对照。