Electrophoresis techniques to investigate defects in oxidative phosphorylation

被引：93

作者：

Calvaruso, Maria Antonietta ^{[1
]}

Smeitink, Jan ^{[1
]}

Nijtmans, Leo ^{[1
]}

机构：

[1] Radboud Univ Nijmegen, Med Ctr, Nijmegen Ctr Mitochondrial Disorders, Dept Pediat,Lab Pediat & Neurol, NL-6500 HB Nijmegen, Netherlands

来源：

METHODS | 2008年 / 46卷 / 04期

关键词：

Blue native electrophoresis; Mitochondria; Oxidative phosphorylation; Two dimensional electrophorsis; Mitochondrial diseases; Enzyme complexes;

D O I：

10.1016/j.ymeth.2008.09.023

中图分类号：

Q5 [生物化学];

学科分类号：

071010 ; 081704 ;

摘要：

Defects in mitochondrial oxidative phosphorylation (OXPHOS) are a frequent cause of severe inherited metabolic disorders and also contribute to aging. The OXPHCS system constitutes five multi-subunit complexes embedded in the mitochondrial inner membrane. Correct function of this system requires proper assembly of the similar to 80 proteins in the complexes, as well as numerous assembly factors. Blue native electrophoresis has become a crucial tool to investigate OXPHOS-related defects in mitochondrial disease patients. In addition, OXPHOS-assembly profiles can be obtained by two dimensional blue native/SDS gel electrophoresis, which provides additional information for identifying disease-causing mutations and insight in the role of specific proteins in the biogenesis of the OXPHOS system. Here we provide a practical guide on how to set-up the basic technique to study OXPHOS defects in patient-derived cells and tissues. (C) 2008 Elsevier Inc. All rights reserved.

引用

页码：281 / 287

页数：7

共 27 条

[21] Identification of mitochondrial complex I assembly intermediates by tracing tagged NDUFS3 demonstrates the entry point of mitochondrial subunits [J].

Vogel, Rutger O. ;

Dieteren, Cindy E. J. ;

van den Heuvel, Lambert P. W. J. ;

Willems, Peter H. G. M. ;

Smeitink, Jan A. M. ;

Koopman, Werner J. H. ;

Nijtmans, Leo G. J. .

JOURNAL OF BIOLOGICAL CHEMISTRY, 2007, 282 (10) :7582-7590

[22] Cytochrome c oxidase subassemblies in fibroblast cultures from patients carrying mutations in COX10, SCO1, or SURF1 [J].

Williams, SL ;

Valnot, I ;

Rustin, P ;

Taanman, JW .

JOURNAL OF BIOLOGICAL CHEMISTRY, 2004, 279 (09) :7462-7469

[23] Advantages and limitations of clear-native PAGE [J].

Wittig, I ;

Schägger, H .

PROTEOMICS, 2005, 5 (17) :4338-4346

[24] Functional assays in high-resolution clear native gels to quantify mitochondrial complexes in human biopsies and cell lines [J].

Wittig, Ilka ;

Carrozzo, Rosalba ;

Santorelli, Filippo M. ;

Schaegger, Hermann .

ELECTROPHORESIS, 2007, 28 (21) :3811-3820

[25] High resolution clear native electrophoresis for In-gel functional assays and fluorescence studies of membrane protein complexes [J].

Wittig, Ilka ;

Karas, Michael ;

Schaegger, Hermann .

MOLECULAR & CELLULAR PROTEOMICS, 2007, 6 (07) :1215-1225

[26] Blue native PAGE [J].

Wittig, Ilka ;

Braun, Hans-Peter ;

Schaegger, Hermann .

NATURE PROTOCOLS, 2006, 1 (01) :418-428

[27] Quantification of muscle mitochondrial oxidative phosphorylation enzymes via histochemical staining of blue native polyacrylamide gels [J].

Zerbetto, E ;

Vergani, L ;

DabbeniSala, F .

ELECTROPHORESIS, 1997, 18 (11) :2059-2064

← 1 2 3 →