Enzymic and physical characterization of diacylglycerol-phosphatidylcholine interactions in bilayers and monolayers

Abstract

The miscibility of 1,3-dioleoylglycerol (DOG) with 1-stearoyl-2-oleoylphosphatidylcholine (SOPC) and 1-palmitoyl-2-oleoylphosphatidylcholine (POPC) dispersed in excess buffer was characterized by physical and enzymatic methods. Thermograms for all SOPC-DOG mixtures exhibit a transition at 5.3 degrees C. Above 0.25 mole fraction of DOG, metastability is observed; after the first scan, a second peak appears at 23.4 degrees C which corresponds to the chain melting of pure DOG. This suggests that a complex or preferred packing array is formed which has a DOG mole fraction of 0.25 (XC). Bilayer morphology is maintained in the metastable state up to 0.8 mole fraction of DOG. Above 0.8, a novel, nonlamellar phase is formed. Fluorescence polarization of 1,6-diphenylhexatriene shows that, relative to SOPC alone, there is little change in the order of the acyl chains up to Xc followed by a large decrease above Xc. Similar results were obtained using POPC. Miscibility was also studied in lipid films at the argon-buffer interface. Isothermal phase diagrams for the mixtures at 15 and 24 degrees C exhibited phosphatidylcholine-DOG complex formation, a region of phosphatidylcholine and complex coexistence, and a region of complex and DOG miscibility. The mole fractions of DOG in the complex (Xc) range from 0.24 to 0.27. Porcine pancreatic phospholipase A2 and pancreatic lipase plus colipase were used as probes of the surface in both the monolayer and bilayer systems. In both systems and with both enzymes, substrate hydrolysis increased abruptly with increasing DOG.(ABSTRACT TRUNCATED AT 250 WORDS)

Regulation of fatty acid oxygen-18 exchange catalyzed by pancreatic carboxylester lipase. 2. Effects of lateral lipid distribution in mixtures with phosphatidylcholine

“…This could indicate a high-order dependence of lipid-protein interaction on the surface concentration of phospholipid. However, for different substrates in films with POPC the critical composition occurs at different mole fractions of substrate and is independent of lipid packing density (Tsujita et al, 1989). Similar independence of the critical composition on lipid concentration for DA/POPC films is indicated by Figure 6.…”

“…Earlier studies had shown that ester substrate/POPC monolayers were miscible in all proportions (Smaby & Brockman, 1985). However, when such films were exposed to a relatively high concentration of CEL (123 nM), a composition-dependent extent of substrate hydrolysis was observed (Bhat & Brockman, 1981;Tsujita et al, 1989). Specifically, in POPC-rich films the extent of hydrolysis was low, generally less than 10% of substrate present, whereas in substrate-rich films hydrolysis was complete.…”

“…However, even if these are eliminated by using monolayers at higher surface pressures with long-chain substrates, nontraditional regulatory effects persist. For example, pancreatic carboxylester lipase (CEL)' activity toward a variety of ester substrates is inhibited in monomolecular films rich in 1 -palmitoyl-2-oleoylphosphatidylcholine (POPC) (Tsujita et al, 1989). POPC reduces enzyme adsorption to these films, but the size of this effect is insufficient to explain the limited extent of substrate hydrolysis (Tsujita et al, 1987;Muderhwa & Brockman, 1990).…”

“…This difference suggested that the lateral organization of the lipids in the interface is an important regulator of enzyme adsorption. In an extension of this argument (Brockman & Muderhwa, 1991), it was proposed that the abrupt change in ester substrate accessibility by surface-bound enzyme at a critical lipid composition (Bhat & Brockman, 1981;Tsujita et al, 1989) reflects a percolation-based transition in lipid head-group organization.…”

Diacylglycerol Kinase from Suspension Cultured Plant Cells

“…However, mixtures of phospholipids and DG were shown to form multilamellar membrane structures (6). Hence, PI and PG may be the natural cofactors that provide DG to the membrane-associated enzyme.…”