生物化学技术

EPR spectroscopy can quantitatively detect free radicals with high specificity and sensitivity in biological samples. However, the natural free radical concentration in biological systems is usually too low for giving sufficient signal intensity which is necessary for EPR. In pr ...

Nitric oxide (NO�), generated by the catalytic action of nitric oxide synthase on L-arginine (1,2), regulates vascular tone, augments cell-cell communications, and governs many intracellular events (3–8). Further, NO� plays a critical role in host immunity, particularly effective aga ...

In recent years, endothelial cells, activated by cytokines (1) have been found to phagocytose Gram-positive and Gram-negative bacteria (2–14), and exhibit microbicidal activity toward S. aureus (15). One of the enzymes induced by cytokine treatment is nitric oxide synthase (NOS), which is ...

Nitric oxide and superoxide rapidly combine to form a toxic reaction product, peroxynitrite anion (ONOO) (1,2). The oxidant reactivity of peroxynitrite is mediated by an intermediate with the biological activity of the hydroxyl radical. However, this product does not appear to be the hydr ...

Aging and the progression of certain degenerative diseases are accompanied by increases in intra- and intercellular fluorescent material, termed lipofuscin and ceroid, respectively (for review, see refs. 1,2. These pigments are observed within granules composed, in part, of damag ...

Liver Slice Technology is a relatively new addition to the battery of in vitro assays of xenobiotic metabolism and toxicity evaluation. As with any developing technology, evaluation and refinement of the technique will be an essential component of its implementation to routine use. Three ...

Thiol groups of cysteine residues in protein have long been known as vulnerable sites, prone to alkylation by electrophiles as well as to oxidation by prooxidant agents, with formation in the latter case of intra- and intermolecular disulfide bridges. A number of studies have shown that such mod ...

The development of controlled-release drug-delivery systems that can achieve an effective concentration for a specific period has been investigated using various drug carriers (1).

The xanthophyll carotenoids lutein and zeaxanthin (see Note 1) are specifically concentrated in the macula of the primate eye, the region of the retina responsible for high-resolution visual acuity necessary for reading, driving, and recognizing faces. They are thought to protect the ma ...

Photodynamic therapy (PDT) is a cancer treatment modality, based on the synergistic photochemical reaction between tumor cells containing accumulated photosensitizer and laser light. The mechanisms of killing of malignant cells were studied by several researchers (1–3). Phot ...

Photodynamic therapy (PDT) is a cancer treatment modality, based on the synergistic photochemical reaction between tumor cells containing accumulated photosensitizer and laser light. The mechanisms of killing of malignant cells were studied by several researchers (1–3). Phot ...

The degradation of misfolded, aged, or no longer needed cytosolic proteins depends largely on the ubiquitin-proteasome system. Proteasomes degrade their substrates into fragments of 3–20 amino acids. Human 20S proteasomes can be purified from human erythrocytes by batch adsorpti ...

Protein ubiquitination is crucial to many diverse and critical functions of cells. Although it has been long known that conjugation of ubiquitin to proteins results in their destruction by the proteasome, recently it has become apparent that reversible protein ubiquitination, part ...

Protocols are given for a variety of techniques used in protein identification of complexes, including identification of in-gel separated proteins and LC-MS/MS. Gels, staining procedures, and peptide extraction protocols that are compatible with mass spectrometry are describ ...

The 20S proteasome is a complex multisubunit protease that is present in all phylae of life. Eukaryotic 26S proteasomes, which are composed of 20S proteasomes and 19S activator complexes, mediate the degradation of ubiquitylated proteins. Biogenesis of proteasomes involves a coordi ...

An important step in the ubiquitin proteolytic cascade is specific recognition of the substrate by a member of the ubiquitin ligases family of proteins—an E3, that is followed by generation of the polyubiquitin degradation signal. For most substrates, it is believed, though it has been demon ...

The fidelity of the protein production process is monitored by quality control machinery, which ensures that aberrant proteins are not deployed throughout the cell. A significant fraction (upwards of 30%) of proteins are degraded by proteasomes shortly after their synthesis, and we have ...

Proteins destined for secretion in eukaryotic cells enter the endoplasmic reticulum (ER) in an unfolded state and are properly folded in this organelle and sent to their final destination. Misfolded or orphan proteins are retained in the ER by a quality control system, retrotranslocated i ...

In this chapter, a genetic approach is presented that leads to the isolation of mutants and to the identification of proteins involved in protein quality control and endoplasmic reticulum-associated degradation (ERAD). The method makes use of a genomic screen of a yeast deletion library (E ...

Components of the ubiquitin-proteasome system function on the surface of the endoplasmic reticulum (ER) to select misfolded proteins for degradation. Herein we describe methods that allow for the study of the pathway for proteasomal degradation of the cystic fibrosis transmembra ...