Enzymatic Reaction (image will open in a new window)
Phospholipase C (originally called lecithinase and also referred to as α-toxin) catalyzes the hydrolysis of the linkage between glycerol and phosphate in lecithin and other phosphatides.
Phospholipase C (PLC) is found in culture media of Clostridium perfringens, Bacillus cereus, Pseudomonas aeruginosa and Staphylococcus aureus (Ottolenghi 1969). PLC is hemolytic, dermonecrotic and lethal. Because it hydrolyzes lecithin of biological membranes it has been found useful in membrane structure studies (O'Toole 1975; Sabban and Loyter 1974; Rottem et al. 1973; Stahl 1973; VanSchijndel et al. 1973). A recent, in-depth report by Low and Saltiel (1988) is of interest.
Other reports on the action of PLC in a number of physiological systems are as follows: Effect on red blood cells and ghosts (Ikezawa and Murata 1964; Coleman et al. 1970; Berengo and Simpkins 1972; DeBoer and Loyter 1971; Lenard and Singer 1968); Effect on muscle preparations (Finean and Martinosi 1965; Martonosi 1968); Inhibition of viral action (Mitzutani and Mitzutani 1964; Freedman and Pastan 1968); Prevention of cholesterol esterification in human serum (Rowen 1964); Effect on glucose transport in free fat cells (Blecher 1965; Rodbell and Jones 1966); Physicochemical studies on the gelatin of hen's egg yolk; Delipidation of yolk plasma by treatment with phospholipase C and extraction with solvents (Kumar and Mahadevan 1970); Effect on nerve membrane (Simpkens et al. 1971); effect on purified myelin (McIlwain and Rapport 1971); Study of bacterial cells (Davie and Brock 1966); Effect on thyroid (Macchia and Pastan 1966); Effect on brain microsomes (Stahl 1973).
Characteristics of Phospholipase C from Cl. perfringens:
Molecular weight: 46,000 ± 500 (Stahl 1973).
Composition: According to Takahashi et al. (1974) there are three major molecular forms separable by isoelectric focusing, having isoelectric points of 5.2, 5.3, and 5.5.
Optimum pH: 7.0 - 7.6.
Activators: Ca2+. Ottolenghi (1967, 1969) reports that the phospholipid substrate must have a net positive charge for enzyme action to occur and that the widely reported Ca2+ activation is a reflection of this requirement. See Renard et al. (1987).
Specificity: Splits lecithin to choline phosphate and diglyceride. It also hydrolyzes sphingomylin, phosphatidylethanolamine and the synthetic water soluble L-α-dicaproyl lecithin. See comparison of possible substrates by Stahl (1973, Table II).
Inhibitors: Phosphonate analogs of glycerophosphatides (Rosenthal and Pousada 1968); ferricyanide (Bangham 1962); bleomycin and polymixin B (Saito et al. 1972); basic proteins such as lysozyme and cytochrome C (Lenaz et al. 1972). Inactivation by EDTA or phenanthroline may be reversed by adding Zn2+ but not Ca2+ (Möllby and Wadstrom 1973).
Stability: Worthington partially purified phospholipase C is stable for at least 5 years when stored at 2 - 8deg.C. Worthington's chromatographically purifed phospholipase is stable at least 6 months when stored at 2 - 8°C.