This investigation examines oxidation conditions under which hemoglobin extracts membrane phospholipid of various headgroup classes and/or acyl chain lengths.  Purified mammalian hemoglobin was oxidized using treatments that result in a nearly quantitative conversion to methemoglobin (ferricyanide) or result in a mixture of methemglobin and the green hemoglobin, choleglobin (hydrogen peroxide).  In red blood cells made echinocytic with exogenous phospholipid, subsequent addition of hemoglobin oxidized with hydrogen peroxide, but not ferricyanide, induced dose-dependent shape reversion to less echinocytic forms, consistent with extraction of phospholipids from the exofacial side of the membrane.  In companion experiments, red cells preloaded with radiolabeled or fluorescently-labeled phosphatidycholine (PC) exhibited greatest extraction of radiolabel or fluorescence signal with exogenous hemoglobin oxidized via hydrogen peroxide but not ferricyanide.  In experiments employing model membranes composed of dimyristoyl-PC liposomes containing radiolabeled PC, phosphatidylserine or phosphatidylethanolamine, subsequent addition of hemoglobin oxidized with hydrogen peroxide extracted radiolabeled lipid with significantly greater efficiency than ferricyanide-treated hemoglobin.  Although radiolabeled lipid extraction was comparable for phospholipids containing saturated acyl chains between twelve and eighteen carbons, radiolabeled lipid extraction of oleoyl-containing phospholipids diminished to levels half that observed with distearoylphosphatidylcholine.  These findings indicate that green hemoglobins such as choleglobin extract lipids from cell membranes regardless of headgroup or acyl chain length, through a process of direct hydrophobic interaction with the membrane surface.

Extraction of phospholipids from human erythrocyte membranes and model membranes by hemoglobin oxidation products

248th National Meeting (2014)

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