Blue Native electrophoresis to study mitochondrial and other protein complexes

被引:312
作者
Nijtmans, LGJ
Henderson, NS
Holt, IJ
机构
[1] Univ Nijmegen, Med Ctr, Nijmegen Ctr Mitochondrial Disorders, NL-6500 HB Nijmegen, Netherlands
[2] SUNY Stony Brook, Dept Pathol, Stony Brook, NY 11794 USA
[3] Wellcome Trust Res Labs, Dunn Human Nutr Unit, Cambridge CB2 2XY, England
基金
英国医学研究理事会;
关键词
Blue Native gel electrophoresis; Blue Native agarose electrophoresis; mitochondria; respiratory chain; oxidative phosphorylation; enzyme complexes; histochemical staining; pulse-chase;
D O I
10.1016/S1046-2023(02)00038-5
中图分类号
Q5 [生物化学];
学科分类号
071010 ; 081704 ;
摘要
The biogenesis and maintenance of mitochondria relies on a sizable number of proteins. Many of these proteins are organized into complexes, which are located in the mitochondrial inner membrane. Blue Native polyacrylamide gel electrophoresis (BN-PAGE) is a method for the isolation of intact protein complexes. Although it was initially used to study mitochondrial respiratory chain enzymes, it can also be applied to other protein complexes. The use of BN-PAGE has increased exponentially over the past few years and new applications have been developed. Here we review how to set up the basic system and outline modifications that can be applied to address specific research questions. Increasing the upper mass limit of complexes that call be separated by BN-PAGE can be achieved by using agarose instead of acrylamide. BN-PAGE can also be used to study assembly of mitochondrial protein complexes. Other applications include in-gel measurements of enzyme activity by histochemical staining and preparative native electrophoresis to isolate a protein complex. Finally, new ways of identifying protein spots in Blue Native gels using mass spectrometry are briefly discussed. (C) 2002 Elsevier Science (USA). All rights reserved.
引用
收藏
页码:327 / 334
页数:8
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