Surface pressure‐dependent cross‐modulation of sphingomyelinase and phospholipase A₂ in monolayers

Abstract

We investigated the ways in which phospholipase A2 and sphingomyelinase are mutually modulated at lipid interfaces. The activity of one enzyme is affected by its own reaction products and by substrates and products of the other enzyme; all this depends differently on the lateral surface pressure. Ceramide inhibits both the sphingomyelinase activity rate and the extent of degradation, and decreases the lag time at all surface pressures. Dilauroyl- and dipalmitoylphosphatidylcholine, the substrates of phospholipase A2 (PLA2), do not affect sphingomyelinase activity. The products of PLA2, palmitic acid and lysopalmitoylphosphatidylcholine, strongly enhance and shift to high surface pressures the activity optimum and the cutoff point of sphingomyelinase. Palmitic acid also shifts to high surface pressures the cut-off point of PLA2 activity. Sphingomyelin strongly inhibits PLA2 at surface pressures above 5 mN/m, while ceramide shifts the cut-off point and the activity optimum to high surface pressures. The sphingolipids increase the lag time of PLA2 at low surface pressures. Both phosphohydrolytic pathways involve different levels of control on precatalytic steps and on the rate of activity that appear independent on specific alterations of molecular packing and surface potential. The mutual lipid-mediated interfacial modulation between both phosphohydrolytic pathways indicates that phospholipid degradation may be self-amplified or dampened depending on subtle changes of surface pressure and composition.

Sphingomyelinase acts by an area-activated mechanism on the liquid-expanded phase of sphingomyelin monolayers

“…This is in keeping with previous work reporting a preference of SMase for the substrate when it is in the fluid state compared to gel/ordered states (15)(16)(17)27). In our previous work (20), we suggested the possibility that the presence of lateral interface could be involved in the completion of the precatalytic steps necessary for the enzyme to reach the steady-state condition (constant rate period).…”

“…Previous studies in both lipid monolayers and bilayers have shown greater activity of SMase when the substrate is in a fluid state than when it is in the gel/ordered state (15)(16)(17). A detailed study of the time course of the SM degradation by SMase showed a correlation between the formation of Cer-enriched LC domains (immersed in a SM-enriched LE continuous phase) and the attainment of full catalytic capacity at the end of a lag period (18,19).…”

The initial surface composition and topography modulate sphingomyelinase-driven sphingomyelin to ceramide conversion in lipid monolayers

“…Recently, enzymaticaly generated Cer-enriched domains were shown to favor apolipoprotein E binding, linking phase segregation events to aterogenesis [9]. To perform their catalytic activity phospholipases such as Phospholipase C, PI-specific Phospholipase C, Phospholipase A 2 and SMase act associated to the membrane/water interface and their enzymatic activity is modulated by interfacial properties of the membrane substrate in a complex manner in which intermolecular packing, phase state, dipole potential, lipid composition, interfacial curvature, among others factors, are involved [10][11][12][13][14][15][16][17][18]. It is known that SMase catalytic activity is highly sensitive to subtle changes of the physicochemical conditions of the lipid interface in both monolayer and bilayer systems [15,[17][18][19][20].…”

Sphingomyelinases and Their Interaction with Membrane Lipids

“…Neither enzyme could produce either aggregation or fusion on its own under those conditions, but when both were added their activities appeared to be mutually potentiated. In a conceptually related study on lipid monolayers at the air-water interface, Fanani and Maggio [146] demonstrated that the activities of SMase and phospholipase A 2 could be mutually modulated. The activity of one enzyme was affected by its own reaction products and by substrates and products of the other enzyme.…”