Phospholipids enzymatic activity

Adaptor Proteins. Figure 1 Adaptor protein domains. A scheme of the domain structures of some well-characterized adaptor proteins is shown. Descriptions of domain characteristics are in main text except C2, binds to phospholipids GTPase activating protein (GAP) domain, inactivates small GTPases such as Ras Hect domain, enzymatic domain of ubiquitin ligases and GUK domain, guanylate kinase domain. For clarity, not all domains contained within these proteins are shown. 

The family of heterotrimeric G proteins is involved in transmembrane signaling in the nervous system, with certain exceptions. The exceptions are instances of synaptic transmission mediated via receptors that contain intrinsic enzymatic activity, such as tyrosine kinase or guanylyl cyclase, or via receptors that form ion channels (see Ch. 10). Heterotrimeric G proteins were first identified, named and characterized by Alfred Gilman, Martin Rodbell and others close to 20 years ago. They consist of three distinct subunits, a, (3 and y. These proteins couple the activation of diverse types of plasmalemma receptor to a variety of intracellular processes. In fact, most types of neurotransmitter and peptide hormone receptor, as well as many cytokine and chemokine receptors, fall into a superfamily of structurally related molecules, termed G-protein-coupled receptors. These receptors are named for the role of G proteins in mediating the varied biological effects of the receptors (see Ch. 10). Consequently, numerous effector proteins are influenced by these heterotrimeric G proteins ion channels adenylyl cyclase phosphodiesterase (PDE) phosphoinositide-specific phospholipase C (PI-PLC), which catalyzes the hydrolysis of phosphatidylinositol 4,5-bisphosphate (PIP2) and phospholipase A2 (PLA2), which catalyzes the hydrolysis of membrane phospholipids to yield arachidonic acid. In addition, these G proteins have been implicated in... 

The question of whether an enzyme is membrane bound or membrane associated is to some extent a matter of semantics. However, it is certainly true that some proteins are readily dissociated from membranes whereas others require quite drastic conditions before they can be dissociated from the membrane. As limiting cases, the former can be designated as membrane associated and the latter as membrane bound. Enzymes that are generally considered membrane bound are firmly embedded in the membrane structure. For example, the mitochondrial coupling factor is strongly coupled to the bilayer structure by hydrophobic polypeptides. The Na+-K+ ATPases that have been purified have a small patch of associated phospholipids when the enzyme is delipidated, enzymatic activity is lost. In fact, membrane-bound enzymes appear to be... 

Lysophospholipids have been found in butter serum by Cho et al. (1977). They characterized the sn-1 and -2 lysophosphatidylcholines and phosphatidylethanolamines. It is not known if these compounds are products of degradation or remnants of biosynthesis. Cho et al. (1977) searched for, but did not find, another possible product of enzymatic degradation of milk, phosphatidic acid. Phosphatidic acid can be formed by the action of phospholipase D on phosphatidylcholine, for example, but this enzymatic activity was not detected. The compound is also an important intermediate in the biosynthesis of lipids, but the concentration in tissue is always very low. The amount is also low in milk. Cho et al. (1977) found 1.2 and 0.9 (percent of total lipid P) of the lyso compounds above. The quantities of the other phospholipids were phosphatidylethanolamine, 27.3 -choline, 29.1 -serine, 13.4 -inositol, 2.5 and sphingomyelin, 25.6. 

Many of the proteins of membranes are enzymes. For example, the entire electron transport system of mitochondria (Chapter 18) is embedded in membranes and a number of highly lipid-soluble enzymes have been isolated. Examples are phosphatidylseiine decarboxylase, which converts phosphatidylserine to phosphatidylethanolamine in biosynthesis of the latter, and isoprenoid alcohol phosphokinase, which participates in bacterial cell wall synthesis (Chapter 20). A number of ectoenzymes are present predominantly on the outsides of cell membranes.329 Enzymes such as phospholipases (Chapter 12), which are present on membrane surfaces, often are relatively inactive when removed from the lipid environment but are active in the presence of phospholipid bilay-ers.330 33 The distribution of lipid chain lengths as well as the cholesterol content of the membrane can affect enzymatic activities.332... 

As an example of an asymmetric membrane integrated protein, the ATP synthetase complex (ATPase from Rhodospirillum Rubrum) was incorporated in liposomes of the polymerizable sulfolipid (22)24). The protein consists of a hydrophobic membrane integrated part (F0) and a water soluble moiety (Ft) carrying the catalytic site of the enzyme. The isolated ATP synthetase complex is almost completely inactive. Activity is substantially increased in the presence of a variety of amphiphiles, such as natural phospholipids and detergents. The presence of a bilayer structure is not a necessary condition for enhanced activity. Using soybean lecithin or diacetylenic sulfolipid (22) the maximal enzymatic activity is obtained at 500 lipid molecules/enzyme molecule. With soybean lecithin, the ATPase activity is increased 8-fold compared to a 5-fold increase in the presence of (22). There is a remarkable difference in ATPase activity depending on the liposome preparation technique (Fig. 41). If ATPase is incorporated in-... 

A large number of macromolecules possess a pronounced amphiphilicity in every repeat unit. Typical examples are synthetic polymers like poly(l-vinylimidazole), poly(JV-isopropylacrylamide), poly(2-ethyl acrylic acid), poly(styrene sulfonate), poly(4-vinylpyridine), methylcellulose, etc. Some of them are shown in Fig. 23. In each repeat unit of such polymers there are hydrophilic (polar) and hydrophobic (nonpolar) atomic groups, which have different affinity to water or other polar solvents. Also, many of the important biopolymers (proteins, polysaccharides, phospholipids) are typical amphiphiles. Moreover, among the synthetic polymers, polyamphiphiles are very close to biological macromolecules in nature and behavior. In principle, they may provide useful analogs of proteins and are important for modeling some fundamental properties and sophisticated functions of biopolymers such as protein folding and enzymatic activity. 

This enzyme is considered to be important to maintenance of the proper level of calcium ion in the erythrocyte. As such it is also considered to be typical of a membrane-bound enzyme. Upon purification of this enzyme from human erythrocytes, it was found that the purified enzyme exhibited no activity until a mixture of a nonionic detergent and a charged phospholipid (e.g., phospha-tidylserine or phosphatidylinositol), were included in the assay system (Nelson and Hanahan, 1985). Ten- to twelvefold increases in activity could be achieved. If a neutral phospholipid such as phosphatidylcholine were substituted for the phosphatidylserine (or phosphatidylinositol), essentially there was little or no enzymatic activity. 

The interaction of certain members of the G protein family with an enzymatic activity known as phospholipase C results in the hydrolysis of the membrane phospholipid, phosphatidyl inositol, into diacylglycerol (DAG) and inosine triphosphate (IP3) shown in Fig. 4.9C (Ross, 1992a Kennedy, 1992 Hardie, 1991). DAG is an activator of the protein kinase C family of enzymes discussed below, and acts by reducing the level of calcium needed for their activation. (Several diacylglycerols, depending on the exact... 

The impact of enzyme activity on the nonhydratable phospholipid content of crude soybean oil was investigated by List et al. (140). Evaluation of flakes subjected to live steam and whole beans treated by microwave heating to inactivate phospholipase D suggests that heat, moisture, and enzyme activity are important factors contributing to the formation of nonhydratable phospholipids in extracted crude oils. Approximately 8-10 minutes of microwave heating is required to completely destroy enzymatic activity. 

Unlike TLL, both the active and inactive forms of PLAf showed periods of immobilization at the layer edge with a much longer residence time for iPLAl. Enzymatic activity is thus not a prerequisite for strong enzyme intercalation at the layer edge, but is clearly a prerequisite for efficient desorption of the enzyme. The products of the hydrolysis reaction cause considerable reorganization and solubilization of the phospholipid bilayer, and either effect could trigger the desorption of enzyme. 

The phospholipid component appears to determine the preferred mode of combination of repeating units with one another. Llpld-free particles polymerize to three-dimensional aggregates, which are essentially bulk phases devoid of enzymatic activity. ReIntroduction of phospholipid restricts the repeating units to "side to side" Interactions Involving predominantly hydrophobic protein-to-protein bonds. This type of interaction affords an enzymatically-active membrane continuum. It has been concluded, therefore, that the essentiality of phospholipid for normal enzymatic function in such systems reflects cheir ability to "direct" membrane formation rather than any specific chemical effect exerted directly on the enzyme. ... 

For lipolytic enzymes—lipases and phospholipases— Braco and co-workers demonstrated that the presence of surfactants (e.g., octyl P-o-glucopyranosides) and phospholipids in the prelyophilization aqueous phase at concentrations above their critical micellar concentrations (i.e., in the form of micelles or vesicles), increased enzymatic activity by several orders of magnitude. It is believed for lipases that the formation of a small bulk aqueous interface induces an open conformation of the enzyme, where a lid that covers the active site recedes. Treatment of crude lipase with 2-propanol may also promote an open conformation of lipases. Similar to bioimprinting, the activation agent, the surfactant, must be rinsed away from the lyo-philized enzyme by anhydrous solvent prior to use. The activity enhancement decreases with increasing water content in the reaction medium. ... 

The evidence to support the idea that many, if not all, membrane proteins possess a immobile, long-lived annulus was of two kinds (a) biochemical and (b) spin-label studies. The first was based upon the fact that to maintain full enzyme activity, at least 30 lipid molecules per protein are required. The second was the presence of an immobile ESR signal at high protein to phospholipid content. Tanford has shown however that phospholipid is not essential for full enzymatic activity and that the hydrophobic fluid environment of a detergent is quite sufficient [75]. Furthermore, deuterium magnetic resonance studies on reconstituted ATPase systems have cast... 

---