Abstract
We prepared highly oriented, multi-lamellar stacks of human red blood cell (RBC) membranes applied on silicon wafers. RBC ghosts were prepared by hemolysis and applied onto functionalized silicon chips and annealed into multi-lamellar RBC membranes. High resolution X-ray diffraction was used to determine the molecular structure of the stacked membranes. We present direct experimental evidence that these RBC membranes consist of nanometer sized domains of integral coiled-coil peptides, as well as liquid ordered (lo) and liquid disordered (ld) lipids. Lamellar spacings, membrane and hydration water layer thicknesses, areas per lipid tail and domain sizes were determined. The common drug aspirin was added to the RBC membranes and found to interact with RBC membranes and preferably partition in the head group region of the lo domain leading to a fluidification of the membranes, i.e., a thinning of the bilayers and an increase in lipid tail spacing. Our results further support current models of RBC membranes as patchy structures and provide unprecedented structural details of the molecular organization in the different domains.
Highlights
When present in the body, aspirin and its metabolites interact with the cyclo-oxygenase (COX) pathway
These red blood cell (RBC) ghosts are applied onto silicon wafers and annealed to form multi-lamellar RBC membrane stacks
X-ray diffraction was used to determine the molecular structure of the RBC membranes
Summary
When present in the body, aspirin (acetylsalicylic acid, ASA) and its metabolites interact with the cyclo-oxygenase (COX) pathway The inhibition of both COX isoforms, COX-1 and COX-2, by higher dose aspirin is believed to lead to analgesic and anti-inflammatory effects, while lower doses, sufficient to inhibit COX-1 activity, lead to anti-platelet activity[7,8]. Place the wafer in the desiccator and apply 100 μL of the sonicated ghosts solution onto the wafer a) Prepare Ghosts: 1. Mix 50 μL RBC solution with 1 mL Buffer in 1.5 mL reaction tubes and place the tubes on ice for 30 minutes. Wash the resulting pellet four times with buffer 6. Place the desiccator in an incubator at 50°C and anneal the membranes for 5 days b) Sonication: 7. Place the desiccator in an incubator at 50°C and anneal the membranes for 5 days b) Sonication: 7. Sonicate this sample 10 times for 5 seconds each
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