Phospholipase injection

The second observation which does not support the unfolded protein model is that when phospholipase A (N. naja venom) was injected into the subphase under the lipid monolayer at equilibrium with globulin, lecithin was readily attacked, as indicated by the rapid fall of surface potential (4, 5, 6). If the penetrated protein were to cover entirely the polar groups of the lipid facing the aqueous subphase (as postulated in the unfolded protein model), the lipid molecules should not be accessible to the lipolytic enzyme. 

Influence of Intermolecular Spacing on Enzymic Hydrolysis of Lecithin Monolayers. When snake venom phospholipase A is injected under a lecithin monolayer, it splits lecithin into lysolecithin and free fatty acid. The change in polar groups of the monolayer results in a change of surface potential. However, if prior to injection of enzyme into the subsolution, a lecithin monolayer is compressed to such a surface pressure that the active site of the enzyme is unable to penetrate the monolayer, hydrolysis does not proceed. For monolayers of dipalmitoyl, egg, soybean, and dioleoyl lecithins the threshold surface pressure values at which hydrolysis does not proceed are 20, 30, 37, and 45 dynes per cm., respectively (40). This is also the same order for area per molecule in their surface pressure-area curves, indicating that enzymic hydrolysis of lecithin monolayers is influenced by the unsaturation of the fatty acyl chains and hence the intermolecular spacing in monolayers (40). 

The action of phospholipase A2 on mixed monolayers of natural and polymerizable lipids can be measured under constant surface pressure by the contraction of the monolayer as a function of time as depicted schematically in Fig. 39. It turns out that the chief parameter influencing the enzymatic activity is the miscibility of the lipid components and not the fact whether the film is polymerized or not. In mixed and demixed membranes the enzyme is able to hydrolyze the natural lipid component, but with considerable differences in the hydrolizing rate (Fig. 40). A pure dilauroyllecithin (DLPC) monolayer is completely hydrolyzed in a few minutes after injecting the enzyme... 

Thwin M. M., Ong W. Y., Fong C. W., Sato K., Kodama K., Farooqui A. A., and Gopalakrishnakone P. (2003). Secretory phospholipase A2 activity in the normal and kainate injected rat brain, and inhibition by a peptide derived from python serum. Exp. Brain Res. 150 427-433. 

St-Gelais F., Menard C., Congar P., Tmdeau L. E., and Massicotte G. (2004). Postsynaptic injection of calcium-independent phospholipase A2 inhibitors selectively increases AMPA receptor-mediated synaptic transmission. Hippocampus 14 319-325. 

Figure 3. Neurophysiological preparation of Trichoplusia ni. Head, thorax and gut are removed. Tungsten electrodes were placed into the hemocoel along side abdominal ganglion VIII (at Rl), the ventral nerve cord (at R2) and the abdominal wall (ground, Rj). Injections of alkaline-dissolved BTI -endotoxin, methamidophos and phospholipase-A2 were into the second pair of abdominal prolegs (Ch). Mechanical sensory stimulation with a glass probe was at the anal proleg (S). Activity in the ventral nerve cord was monitored through 24 h post-treatment (38-40) (see Figure 4).

Figure 4. Time dependency of nervous activity in the ventral nerve cord of Trichoplusia ni injected with 3.7 PPM alkaline-dissolved BTI 6-endotoxin (Sandoz), with 10 PPM methamidophos (MMP) and with 35 PPM phospholipase-A2 (P-A2). Mechanical sensory stimulation is given at arrow S. The control response was the same as the recording for P-A2. BTI and P-A2 were injected into T ni at their respective LDjq.

The crystal is only one of the toxins produced by these bacteria. In 1953 Toumanoff (28) reported on a study of the genus Bacillus and their phospholipase C activity. In 1954 he found (29) that the precipitate of this enzyme from broth filtrates caused death when injected or fed to larvae of the greater wax moth [Galleria mellonella (L.)] In 1955 Heimpel (13) showed that strains of B. cereus and B. thuringiensis... 

The ll-palm-A -THC can be hydrolyzed to II-OH-A -thc by cholesterol esterase and triacylglycerol lipase but not by phospholipase A, acetylesterase or phosphotransacetylase (16). An attempt to modify the retention of fatty acid-conjugated DDT metabolites was carried out by injecting the DDT-treated rats with sodium salt of various bile acids, heparin or lecithin of which all were known to affect the esterification or ester hydrolysis by the cholesterol esterase system. The results Indicated a significant decrease in the retention of the conjugated DDT metabolites in the rat liver and spleen (17). 

Bravo-Cuellar A, Homo-Delarche F, Ramos-Zepeda R, Dubouch P, Cabannes J, Orbach-Arbouys S. Increased phagocytic activity of peripheral blood monocytes after intravenous injection of phospholipase A2 to monkeys. Immunol Lett 1991 28 5-10. 

The high substrate specificity of some enzymes can be used as the basis for highly selective determinations of organic phosphorus species. For example, Amini (2001) determined phosphatidyl choline using coimmobilized phospholipase C, alkaline phosphatase and choline oxidase (Fig. 1.2a). A stoichiometric product of this sequence is hydrogen peroxide, and this was determined using the luminol chemiluminescence reaction, using the flow injection manifold shown (Fig. 1.2b) to provide rapid, sensitive and reproducible indirect measurement of phosphatidyl choline (Amini,... 

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