Ordered-disordered domain coexistence in ternary lipid monolayers activates sphingomyelinase by clearing ceramide from the active phase

ELISA CARMEN ALE; BRUNO MAGGIO; MARIA L. FANANI

BIOCHIMICA ET BIOPHYSICA ACTA-BIOMEMBRANES

ELSEVIER SCIENCE BV

Lugar: Amsterdam; Año: 2012 vol. 1818 p. 2767 - 2776

0005-2736

We explored the action of sphingomyelinase (SMase) on ternary monolayers containing phosphatidylcholine, sphingomyelin (SM) and dihydrocholesterol, varying along a single tie line of phase coexistence. SMase activity exhibited higher rates and extents of hydrolysis when the film is within the liquid-expanded (LE)/liquid-ordered (LO) coexistence range, compared to monolayers in the full LO phase. Since Alexa-SMase preferably adsorbs to the LE phase, and no direct correlation was found between enzymatic activity and domain borders, we postulate that the LE phase is the active phase for ceramide (Cer) generation. The enzymatically generated Cer organizes in different manners depending on the initial LE/LO ratio. The action of SMase in Chol-poor monolayers leads to the formation of Cer-enriched domains, while in Chol-rich monolayers it results in the incorporation of Cer in the LO phase and formation of new Chol- and Cer-enriched domains. The novel mechanism proposed here provides explanation for the favored action of SMase on interfaces that exhibit LE-LO phase coexistence: the LO phase sequesters the product Cer causing its depletion from the more enzyme-susceptible LE phase, thus decreasing reaction product inhibition. Furthermore, LO domains function as substrate reservoir by allowing rapid exchange of the substrate from this phase to the SM-depleted LE phase.