Effect of sphingosine and stearylamine on the interaction of phosphatidylserine with calcium. A study using DSC, FT-IR and 45Ca 2+-binding

Abstract

The lamellar gel to lamellar liquid-crystalline phase transition of dipalmitoylphosphatidylserine (DPPS) multilamellar membranes is abolished by the presence of Ca 2+ at DPPS/Ca 2+ molar ratios of 2:1 or lower. However, when equimolar sphingosine (SPH) or stearylamine (SA), which are positively charged at the pH studied in this work, were included in DPPS vesicles, the phase transition of DPPS was still observed by differential scanning calorimetry, even in the presence of very high Ca 2+ concentrations such as a DPPS/Ca 2+ molar ratio of 1:10. According to that, ΔH was similar for samples formed by equimolar DPPS and SPH and SA, either in the presence or in the absence of Ca 2+, whereas no phase transition was observed for the pure phospholipid in the presence of Ca 2+ at molar ratios lower than DPPS/Ca 2+2:1. 45Ca 2+-binding experiments showed that for DPPS/SPH or DPPS/SA molar ratios of 2:1, only half of the Ca 2+ was bound to DPPS with respect to pure DPPS, i.e., in the absence of SPH or SA. At concentrations of SPH or SA equimolar with DPPS, the Ca 2+ binding was nearly abolished. The effect of SPH and SA on the apparent p K app of the carboxyl group f of DPPS was also studied in the presence and in the absence of Ca 2+ by using Fourier transform infrared spectroscopy. The dehydration of the phosphate group of DPPS induced by the binding of Ca 2+ was followed through the observation of the PO 2 − antisymmetric stretching, and the percentage of dehydrated PO 2 − groups quantitatively assayed. It was again confirmed that, in the presence of equimolar concentrations of SPH or SA, Ca 2+, at concentrations which are saturating for pure DPPS, was not bound at all to DPPS. It was also found that the p K app was considerably shifted to lower values in the presence of the amino bases, decreasing from 4.6 in pure DPPS to 2.1 and 2.2 for the equimolar mixtures of DPPS with SPH and SA, respectively. These results show that SPH and SA, being positively charged molecules anchored in the membrane, are able of preventing the binding of positively charged ions such as Ca 2+ through an electrostatic charge neutralization.