Calcium mediated interaction of calf-thymus DNA with monolayers of distearoylphosphatidylcholine: a neutron and X-ray reflectivity study

Aleksandra P Dabkowska,Jonathan P Talbot,David J Barlow,M Jayne Lawrence,John R P Webster,Andrew Nelson,Leide Cavalcanti,Giovanna Fragneto

doi:10.1039/c3sm50350j

Abstract

X-ray and neutron reflection studies, the latter in conjunction with contrast variation, have been combined to study the interaction of calf thymus DNA (ctDNA) with monolayers of distearoylphosphatidylcholine (DSPC) in the presence of 20 mM Ca2+ ions, at the air–liquid interface as a function of surface pressure (10, 20, 30 and 40 mN m−1). Analysis of the X-ray and neutron reflection data showed that, regardless of the surface pressure of the monolayer, a layer of ctDNA was present below the DSPC lipid head groups and that this ctDNA-containing layer (thickness ∼12.5 to 15 A) was separated from the DSPC head groups by a layer of water of ∼9 A thickness. The thickness of the ctDNA-containing layer was thinner than that reported for monolayers of cationic lipid at the air–water interface (18–25 A) although in these monolayers no water layer separating the lipid head groups from the layer containing ctDNA has been reported. At all surface pressures the amount of ctDNA present in the layer was in the range 30–40% by volume. As no significant re-arrangement of the DSPC film was required to accommodate the presence of the ctDNA, this suggests that the distribution of charges in the lipid film matches well the charge spacing of ctDNA. Brewster angle microscopy measurements of DSPC on water in the absence of Ca2+ showed the presence of a continuous film containing small, regular shaped domains at all four surface pressures examined. When Ca2+ ions were present in the sub-phase, although the film was still continuous, the domains comprising the film were more irregular in appearance while the presence of Ca2+ ions and ctDNA resulted in the domains becoming smaller and more regularly packed on the surface.

Highlights

The ability to arti cially engineer genes means that it is possible to tackle diseases hitherto inaccessible to traditional means of drug treatment
In agreement with others,[17] the p–A0 isotherm obtained for a monolayer of h-DSPC on H2O at 295 Æ 2 K in the present study is consistent with that of a gel phase, Lb, lipid over the whole of the range of surface pressures up to the monolayer collapse point, which occurred at a surface pressure of $60 mN mÀ1
If the isotherm is extrapolated to its collapse point, a limiting interfacial area per molecule of $42 A2 is obtained, which is precisely what would be expected given that the limiting cross-sectional area for each of the molecules' two octadecyl chains would be 21 A2, further supporting the hypothesis that DSPC is in the gel phase under the conditions of the experiment

Summary

Introduction

The ability to arti cially engineer genes means that it is possible to tackle diseases hitherto inaccessible to traditional means of drug treatment. The p–A0 isotherms of the other ‘contrasts’ used in the neutron re ectivity study, namely d70-DSPC on water and acmw, d83-DSPC on acmw and h-DSPC on D2O, were determined (data not shown) both to check the purity of the various isotopic forms of the lipid and to verify that deuteration of either the DSPC molecule or the aqueous sub-phase did not cause any signi cant alteration in surface behaviour – a result which is important for the subsequent X-ray and neutron re ection studies.

Results

Conclusion