Blue native polyacrylamide gel electrophoresis (BN-PAGE) employs the dye Coomassie for the labeling of proteins and protein complexes under native conditions. Electrophoresis under native conditions subsequently allows resolution of proteins and protein complexes acc ...
Iron-sulfur (Fe-S) clusters are cofactors of many proteins that are involved in central biochemical pathways, such as oxidative phosphorylation, photosynthesis, and amino acid biosynthesis. The assembly of these cofactors and the maturation of Fe-S proteins require complex cell ...
Mitochondria are dynamic organelles with activities that adjust to altering physiological conditions and variable metabolic demands. A conserved proteolytic system present within the organelle exerts essential functions during the biogenesis of mitochondria and ensu ...
This chapter describes a luciferase-based protocol to measure adenosine triphosphate (ATP) production in isolated mitochondria of Trypanosoma brucei. The assay represents an excellent method to characterize the functionality of isolated mitochondria. Comparing the ATP p ...
The adenosine triphosphate (ATP) synthase (F1-F0 complex) of the mitochondrial inner membrane is responsible for making nearly all of the ATP utilized by eukaryotic organisms. The enzyme is an oligomer of more than 20 different subunits, 14 of which are essential for its catalytic activity. T ...
Mitochondria not only are a source of reactive oxygen species (ROS) but also are sites of oxidative damage. In plants, mitochondria must normally operate when there are high levels of ROS produced during photosynthesis and photorespiration. These levels are further enhanced during biot ...
Mitochondria are key players in cell death following intrinsic and, in some cell types, extrinsic stimuli. The recruitment of the mitochondrial pathway results in mitochondrial dysfunction and release of intermembrane space proteins like cytochrome-c that are required in the cyto ...
Mitochondria play a vital role in the regulation of intracellular calcium dynamics. Fluorescent dyes can be used to provide a direct measurement of the redox state, mitochondrial membrane potential, and mitochondrial calcium content. The simplicity of this approach lends itself to hi ...
The field of mitochondrial dynamics has received a great deal of attention as a result of a number of studies linking mitochondrial fission and fusion machinery to apoptosis. Specifically, elevated levels of mitochondrial fission or compromised mitochondrial fusion can sensitize c ...
The budding yeast Saccharomyces cerevisiae has many advantages as a model system, but until recently high-resolution microscopy was not often attempted in this organism. Its small size, rounded shape, and rigid cell wall were obstacles to exploring the cell biology of this model eukaryote. ...
Electron microscopy (EM) has been a central tool in delineating the subcellular organization and function of the eukaryotic cell. It has provided valuable information on the organization of the Golgi complex; the polarized distribution of proteins on the plasma membrane; and fundamen ...
The study of mitochondrial division proteins has largely focused on yeast and mammalian cells. We describe methods to use Caenorhabditis elegans as an alternative model for studying mitochondrial division, taking advantage of the many wonderful resources provided by the C. elegans c ...
Although 30 years ago it was strongly suggested that some cytoplasmic ribosomes are bound to the surface of yeast mitochondria, the mechanisms and the raison d’ětre of this process are not understood. For instance, it is not perfectly known which of the several hundred nuclearly encoded genes ha ...
Because virtually all cellular processes are based on proteins, detailed knowledge of the mitochondrial proteome represents an integral part of understanding mitochondrial function. The analysis of very complex protein mixtures such as entire cell organelles makes high deman ...
Transcriptomic analysis using quantitative reverse transcriptase polymerase chain reaction (QRT-PCR) facilitates analysis of nuclear and mitochondrial-encoded mitochondrial genes, enabling mechanisms and regulation of signaling pathways to be explored. To illu ...
Significant efforts have sought to uncover the protein profile of Arabidopsis mitochondria to act as a model for the mitochondrial proteome from plants. A combination of techniques have been undertaken to achieve this goal. We outline a basic two-dimensional polyacrylamide gel elect ...
The impact of mitochondria on several fundamental cellular processes is reflected in their involvement in the pathophysiology of common diseases such as Parkinson’s disease, diabetes, and obesity and a wide range of monogenic disorders primarily associated with energy impairme ...
The insulin-like growth factor-binding proteins (IGFBPs) are carrier proteins that are present in all extracellular fluids (1). Similarly, insulin-like growth factor-I (IGF-I) and -II are synthesized by connective tissue cells that are present within the stroma of all tissues, and the ...
Recent advances in developmental biology have highlighted the importance of the synergistic relationships between structural components of the extracellular matrix (ECM) and growth factors that play fundamental roles in tissue morphogenesis and repair. Increasing evide ...
Dissociated cell culture has proved extremely valuable for studying the functional relationship between specific cell surface and extracellular matrix (ECM) molecules and a variety of cellular behaviors such as proliferation, differentiation, migration, and neuronal pro ...
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