生物化学技术

Studying protein–protein interactions is critical to our understanding of the signaling pathways. The Telomere Interactome is assembled around the telomeres and consists of proteins and factors from diverse pathways. Dissecting how this protein network contributes to telom ...

Telomeres, the specialized DNA-protein complexes found at the termini of all linear eukaryotic �chromosomes, protect chromosomes from degradation and end-to-end fusion. The protection of telomeres 1 (POT1) protein binds the single-stranded overhang at the ends of chromosomes in d ...

Telomere Chromatin Immunoprecipitation (ChIP) is an experimental method used to determine whether proteins are associated with telomere DNA inside the nucleus of cells or tissues. Telomere-associated proteins are covalently cross-linked to telomere DNA, and then immunopre ...

Telomere dysfunctions, rendered through replicative attrition of telomeric DNA or due to the inhibition of shelterin components, are recognized as DNA double-stranded breaks (DSBs) by the DNA damage repair (DDR) pathway. This leads to the activation of DNA damage checkpoint sensors, i ...

Dysfunctional telomeres arising either through natural attrition due to telomerase deficiency or by the removal of telomere binding proteins are recognized as double stranded breaks (DSBs). Repair of DSBs is crucial for the maintenance of genome stability. In mammals, DSBs are repair ...

Human telomerase is a ribonucleoprotein (RNP) that synthesizes DNA repeats at the ends of chromosomes and maintains telomere length and genome stability. The enzyme comprises telomerase RNA (hTR) (which provides the template for telomere addition) and several protein subunits, in ...

Fluorescence in situ hybridization (FISH) has become a powerful tool for exploring genomes at the level of chromosomes. The procedure can be used to identify individual chromosomes, rearrangements between chromosomes, and the location within a chromosome of specific DNA sequences s ...

Levels of telomerase activity can be an indicator of the proliferative potential of somatic cells and may serve as a diagnostic biomarker of malignancy. Telomeric repeat amplification protocol (TRAP) is a fast and sensitive PCR-based assay for detection and measurement of telomerase a ...

Immortalised cultured cells are powerful tools to assess the function of ectopically expressed proteins. However, it must be ensured that the protein of interest is functional in the host system and display native behaviour. In particular, mitochondrial uncoupling proteins (UCPs) d ...

The concentration of calcium ions in the mitochondria has a profound impact on mitochondrial function, modulating respiratory activity at physiological concentrations, while causing lethal damage during calcium overload. The “rhod” series of calcium sensitive fluorescent ...

Ca2+-sensitive electrode as a practical approach is used to follow Ca2+ changes in the medium and particularly useful to study mitochondrial Ca2+ uptake (or release); this method permits the continuous recording of Ca2+ movements through the mitochondrial inner membrane. In this chapte ...

Mitochondria are considered as the main source of reactive oxygen species (ROS) in the cell. For this reason, they have been recognized as a source of various pathological conditions as well as aging. Chronic increase in the rate of ROS production is responsible for the accumulation of ROS-assoc ...

Oxidative phosphorylation is an important energy-conserving mechanism coupling mitochondrial electron transfer to ATP synthesis. Coupling between respiration and phosphorylation is not fully efficient due to proton and electron leaks. In this chapter, methods are presen ...

Opening of a large conductance channel in the inner mitochondrial membrane, known as the mitochondrial permeability transition (MPT) pore, has been shown to be a primary mediator of cell death in the heart subjected to ischemia-reperfusion injury. Inhibitors of the MPT have been shown to red ...

Mitochondrial energy metabolism depends upon high-flux and low-flux electron transfer pathways. The former provide the energy to support chemiosmotic coupling for oxidative phosphorylation. The latter provide mechanisms for signaling and control of mitochondrial funct ...

Laser scanning confocal microscopy provides the ability to image submicron sections in living cells and tissues. In conjunction with pH-indicating fluorescent probes, confocal microscopy can be used to visualize the distribution of pH inside living cells. Here, we describe a confoc ...

Bioenergetic Science started in seventh century with the pioneer works by Joseph Priestley and Antoine Lavoisier on photosynthesis and respiration, respectively. New developments were implemented by Pasteur in 1860s with the description of fermentations associated to micro ...

Nuclear magnetic resonance (NMR) spectroscopy is a technique with an increasing importance in the study of metabolic diseases. Its initial important role in the determination of chemical structures (1, 2) has been considerably overcome by its potential for the in vivo study of metabolism ( ...

The advent of techniques with the ability to scan massive changes in cellular makeup (genomics, proteomics, etc.) has revealed the compelling need for analytical methods to interpret and make sense of those changes. Computational models built on sound physico-chemical mechanistic ba ...

Protocols for high-resolution respirometry (HRR) of intact cells, permeabilized cells, and permeabilized muscle fibers offer sensitive diagnostic tests of integrated mitochondrial function using standard cell culture techniques and small needle biopsies of muscle. Mul ...