The 4,5-double bond of ceramide regulates its dipole potential, elastic properties, and packing behavior

Abstract

The biological activities of ceramides show a large variation with small changes in molecular structure. To help understand how the structure regulates the activity of this important lipid second messenger, we investigated the interfacial features of a series of synthetic ceramide analogs in monomolecular films at the argon-buffer interface. To minimize differences arising from the N-acyl moiety, each analog had either a N-hexadecanoyl or a N-cis-4-hexadecenoyl moiety amide linked to the nitrogen of the sphingosine backbone. We found that the trans 4,5-unsaturation in the sphingosine backbone promoted closer packing and lower compressibilities of ceramide analogs in interfaces relative to comparable saturated species. Moreover, structures with this feature exhibited dipole potentials as much as 150-250 mV higher than comparable compounds lacking 4,5-unsaturation. The results support the hypothesis by M.C. Yappert and co-workers that trans unsaturation in the vicinity of C4 of the sphingoid backbone augments intramolecular hydration/hydrogen bonding in the polar region. This intramolecular hydration may allow the close packing of the ceramide molecules and engender their high dipole potentials. These properties of ceramides and their analogs may be important determinants of biological function.

FIGURE 1

The structures and codes of d-erythro-ceramide and its analogs used in this study.

FIGURE 2

Role of C4 unsaturation in regulating the surface behavior of ceramide analogs. (A) Surface pressure-molecular isotherms and (B) dipole potential-molecular area isotherms for Δ4^t(2N)16:0 (▪), ΔΔ4(2N)16:0 (•), [DE](2N)16:0 (▴), [LT](2N)16:0 (▾), and Δ4Δ5(2N)16:0 (♦).

FIGURE 3

Role of trans double bond position in the sphingoid base in regulating surface behavior of ceramide analogs. (A) Surface pressure-molecular area isotherms and (B) surface potential-molecular area isotherms for Δ15^t(2N)16:0 (▪), Δ7^t(2N)16:0 (•), Δ5^t(2N)16:0 (▴), and Δ4^t(2N)16:0 (▾).

FIGURE 4

Comparison of the surface behavior of N-4-cis-hexadecenoyl- and N-hexadecanoylceramide analogs. (A–C) Surface pressure-molecular area isotherms and (D–F) surface potential-molecular area isotherms for (A and D) Δ4^t(2N)16:1 and Δ4^t(2N)16:0; (B and E) Δ5^t(2N)16:0 and Δ5^t(2N)16:1; and (C and F) Δ15^t (2N)16:0 and Δ15^t(2N)16:1. Lines without symbols denote the N-hexadecanoyl species.

FIGURE 5

Role of polar headgroup structure in regulating surface behavior of ceramide analogs. (A) Surface pressure-molecular isotherms and (B) surface potential-molecular area isotherms for ΔΔ4(3N)16:0 (▪), Δ4^c(3N)16:0 (•), 4OH(2N)16:0 (▴), and Δ3^t5OH(2N)16:0 (▾).

FIGURE 6

Dominance of the sphingoid base double bond position in determining the dipole potential at 35 mN/m N-acyl saturated and unsaturated ceramide analogs; N-hexadecanoylceramides (▪), N-4-cis-hexadecenoylceramides (▴), and dihydroceramide (·······).