细胞技术

There is now increasing evidence that phospholipase D (PLD) activity can be stimulated by a range of hormones, growth factors, and neurotransmitters in a range of cell types (1). The enzyme generally catalyzes the hydrolysis of phosphatidylcholine (PtdCho) to produce phosphatidate (Pt ...

The serine/threonine kinase protein kinase C (PKC) is regarded as playing a key role in stimulation of cellular responses in many different cells and tissues. There are now known to be at least 10 different isoenzymes of PKC (α, βI, βII, γ, δ, ɛ,ζ, η, θ, and λ @#@) which vary in their cofactor requirement for activati ...

The enzyme Ca2+/calmodulin-stimulated protein kinase II (CaM-PK II) belongs to a family of calmodulin-stimulated serine/threonine protein kinases that are widely distributed in nature, but are especially enriched in the brain (1–3). CaM-PK II is also found in a number of subcellular frac ...

Many cell-surface receptors via G-proteins activate phosphoinositidase C, which catalyzes the hydrolysis of phosphatidylinositol 4,5-bis-phosphate to produce the second messengers, myo-inositol 1,4,5-trisphosphate and diacylglycerol (1). Ins(1,4,5)P3 interacts w ...

The early methods for calcium measurement involved microinjection of calcium-sensitive proteins, such as aequorin or obelin, into large cells (1, 2) or the use of microelectrodes (3). Both techniques are still employed, however, with much improved sensitivity allowing investigati ...

Diacylglycerol (DAG) found within cells is derived from two broad sources. First, in the de novo biosynthetic pathway, glycerol 3-phosphate is converted by two acylation steps to phosphatidic acid (PA), which is converted to DAG by phosphatidate phosphohydrolase (PPH). This is part of the pa ...

Phosphoinositide hydrolysis by phospholipase C (PLC) is a widespread transduction mechanism by which activation of many neurotransmitter and hormone receptors triggers the formation of the second messengers inositol 1,4,5-trisphosphate and 1,2-diacyl-glycerol (1–3). Co ...

Excitatory amino acid receptors are the predominant type of neurotransmitter receptor in the vertebrate central nervous system (CNS). They are membrane-spanning proteins that mediate the stimulatory actions of glutamate and possibly other related endogenous amino acids. Exc ...

The phosphoinositide cycle is now established as a major pathway by which a variety of hormones, neurotransmitters, and other signaling molecules can generate second messengers and thus affect the intracellular milieu (1). Receptor-mediated activation of membrane-associated ...

It has now been established that agonist stimulation of a large variety of cell-surface receptors promotes hydrolysis of inositol phospholipids through activation of a phosphoinositide-specific phospholipase C (1, 2). Phosphodiesteric cleavage of phosphatidylinositol ...

The ubiquitous intracellular second messengers 3′,5′-cyclic adenosine monophosphate (cAMP) and 3′,5′-cyclic guanosine monophosphate (cGMP) mediate the effects of a large variety of hormones and neurotransmitters. Both of these nucleotides are inactivated by a large group of en ...

Cyclic GMP (cGMP)-dependent protein kinase (G kinase) can be purified from bovine lung with the combination of ion-exchange chromatography, ammonium sulfate precipitation, and cyclic nucleotide affinity chromatography (1). In this chapter, we describe the purification proce ...

A multitude of natural and pharmacological agents, including hormones, neurotransmitters, and phosphodiesterase inhibitors, raise intracellular cAMP levels and, in the vast majority of instances, the physiological effects are the result of activation of cAMP-dependent pr ...

The recent advances in the understanding of the mechanism of action of molecules like nitric oxide and natriuretic peptides have increased the interest in cyclic GMP (cGMP) metabolism. Several techniques have been described to measure cGMP levels that include: 1. A binding assay

Radioligand binding is an extremely versatile technique that can be applied to a wide range of receptors in a variety of preparations, including purified and solubilized receptors, membrane preparations, whole cells, tissue slices, and even whole animals. The basic method is very easy to pe ...

Plasma membranes encompass a complex and varying set of proteins essential to life. In addition, plasma membrane proteins represent the majority of all known drug targets. The characterization of plasma membrane proteomes is, therefore, of eminent importance. A current bottleneck is t ...

A full understanding of leukocyte responses to external stimuli requires knowledge of the full complement of proteins found on their surfaces. Systematic examination of the mammalian cell surfaces at the protein level is hampered by technical difficulties associated with proteo ...

Defining membrane proteomes is fundamental to our understanding of many physiological and pathophysiological processes. Their separation and identification is hence a key issue in basic and biomedical research. Due to their hydrophobic character, few high-resolution techn ...

Platelets (thrombocytes) are the smallest human blood cells and are pivotal in processes of hemostasis and thrombosis. Central to their function, the activation of platelets includes a complex interplay of adhesion and signalling molecules mediated via the plasma and inner membrane. ...

Membranous vesicles are constitutively released by a multitude of cell types. Following fusion of multivesicular bodies with the plasma membrane, endocytic vesicles, 30–90nm in size termed exosomes are released extracellularly. Whilst several groups have reported the presen ...