细胞技术

Coated vesicles are implicated in a number of receptor-mediated transport events within eukaryotic cells. In particular, they are required for the export of newly synthesized lysosomal enzymes from the trans-Golgi network, and at the plasma membrane they are responsible for receptor ...

Receptor-mediated endocytosis is an important mechanism by which the cell is able to take in macromolecules from the external environment and carry them through the necessary metabolic processes. Studies of the sequential steps of these events require methods of following the fate of the ...

The plasma membrane (PM) of polarized epithelial cells is composed of two physically continuous, but compositionally and functionally distinct domains: basolateral and apical. In hepatocytes, the basolateral domain includes the sinusoidal front, which faces the space of Disse and ...

The membrane organelles of an organized tissue, such as rat liver, that will always form closed vesicles, irrespective of the mode of homogenization, are the tubular membranes of the smooth and rough endoplasmic reticulum. Almost without exception, these vesicles retain their normal ori ...

Continuous-flow electrophoresis (CFE), which separates particles on the basis of surface charge density, should be regarded as an adjunct to centrifugation rather than as an alternative for membrane fractionation. The equipment is expensive and has been used for relatively specia ...

This fraction is defined broadly as the material that will sediment at about 3500g for 10 min from a postnuclear supernatant. It contains the nuclei that failed to sediment at 1000g for 5–10 min, the largest mitochondria, and very few other organelles, such as lysosomes and peroxisomes.

Of all the organdies in rat hepatocytes, the nucleus is the largest and the most dense. It is therefore relatively easy to isolate in high purity and high yield. The majority of methods (1,2) involve the homogenization of the liver in isoosmotic sucrose followed by differential centrifugation and ...

Glycolipids are an important group of diverse molecules present in the plasma membrane and to a more limited extent in the intracellular membranes of the Golgi and lysosomal systems of eukaryotic cells. They are prominent as antigenic determinants and as the ligands for specific lectin bin ...

The function of many glycoproteins is directly related to the structure of their oligosaccharide chains (1–5). Subtle changes in oligosaccharide structure involving the linkage of or loss of single monosaccharide units are known to occur in many biological events. Knowledge of this fine ...

A strategy for determining the structure of a glycosaminoglycan (GAG) can be divided into three areas. The first is to establish its saccharide composition. This requires complete depolymerization of the parent chains to disaccharide products that can be purified, separated, and iden ...

The proteoglycans (PGs) are a large and varied family of complex macromolecules whose physical properties are dominated by their large sulfated polysaccharide chains (for a recent review, see ref. 1). They are widely distributed in the animal kingdom, but are not associated with any particu ...

Membrane enzymes may be peripheral or integral proteins. The attachment of peripheral enzymes is mediated mainly by association with other membrane proteins and consists primarily of hydrophilic, electrostatic interactions. Integral membrane proteins are primarily bound ...

The high resolution capacity of polyacrylamide gel electrophoresis (PAGE) (see Chapter 19) has resulted in the widespread application of this group of techniques to protein separations. PAGE procedures can provide characterization of proteins in terms of their charge, size, relat ...

As discussed in other chapters in this book (see especially Chapters 6, 7, 9, 23, and 24), many nucleases were first identified as such by their sequence identity to known nucleases. Several of these were isolated because they exhibited a seemingly unrelated activity (protein kinase, angiogene ...

Restriction endonucleases coupled with DNA methyltransferases form the restriction-modification (RM) systems that occur ubiquitously among bacteria. They protect bacterial cells against bacteriophage infection by cleaving incoming foreign DNA highly specifica ...

Pulmozyme� recombinant human deoxyribonuclease I (DNase I) is currently used as a therapeutic for cystic fibrosis (CF) (1) and may be effective in the treatment of systemic lupus erythematosus (SLE) (2,3). As described in Chapter 20, degradation of high-molecular-weight DNA following in ...

Human deoxyribonuclease (DNase I) is an important clinical agent currently used in the treatment of cystic fibrosis (CF) patients (1). It is inhaled into the airways, where it degrades DNA to lower molecular weight fragments, thus reducing the viscoelasticity of CF sputum and improving lung f ...

Reverse transcription is the process whereby the single-stranded RNA genome of a retrovirus is converted into double-stranded DNA by the DNA polymerase and ribonuclease H (RNase H) activities of virus-coded reverse transcriptase (RT) (2,16). The former activity of reverse transcrip ...

Ribonuclease A (RNase A), isolated from bovine pancreas, maintains a well-deserved place in the history of modern biochemistry, as many of the earliest studies on amino-acid sequencing, protein crystallography, and protein folding were performed on this stable and abundant protein (1). ...

Lectins, proteins that specifically bind carbohydrates, are widely distributed in animal and plant species. Functions of animal lectins include uptake of serum asialo-glycoproteins into liver, self-defense mechanisms, and modulation of cell-cell interactions during diff ...