Interaction of phospholipid bilayers and monolayers with nominally soluble proteins

Membranes are important cellular structures fulfilling the biological role of compartmentalization. Many types of molecules show important interactions with cell membranes. Previous work has suggested that a cytosolic protein, glyceraldehyde-3-phosphate dehydrogenase, is involved in the maintenance of red blood cell shape. The studies presented in this thesis explore the interaction of nominally soluble proteins with model membrane systems.; A method of preparing a homogeneous population of small unilamellar lipid vesicles is presented. The size of vesicles prepared by this method was confirmed by dynamic light scattering.; The interactions of carbonmonoxyhemoglobin, glyceraldehyde-3-phosphate dehydrogenase, and polyhistidine with vesicles at physiological pH were studied using NMR spectroscopy. Addition of carbonmonoxyhemoglobin to dimyristoylphosphatidylcholine:dimyristoylphosphatidylserine vesicles results in upfield chemical shifts of lipid acyl chain proton resonances and line broadening of all acyl proton peaks. 31P NMR spectra show line broadening of the phosphatidylserine resonance. Some titratable histidine residues show NOE interactions with lipid resonances. These results are consistent with heme dissociation from carbonmonoxyhemoglobin followed by heme intercalation into the lipid bilayer. Addition of glyceraldehyde-3-phosphate dehydrogenase to vesicles also results in line broadening of lipid resonances. Polyhistidine causes general line broadening upon addition to vesicles.; The interactions of carbonmonoxyhemoglobin, glyceraldehyde-3-phosphate dehydrogenase, and polyhistidine with lipid monolayers at the air-water interface were studied. Carbonmonoxyhemoglobin and glyceraldehyde-3-phosphate dehydrogenase both interact extensively with monolayers containing lipids with negatively charged headgroups. This interaction decreases with increasing pH. Carbonmonoxyhemoglobin interacts more strongly with monolayers at low pH, while the glyceraldehyde-3-phosphate dehydrogenase-monolayer interaction shows more complex behavior. These interactions can be modeled as diffusion-limited processes, with kinetic data fit to a stretched exponential equation. The significance of these kinetics are discussed. Polyhistidine interacts only with monolayers containing lipids with negatively charged headgroups.; The combination of light scattering, NMR, and monolayer results indicates that the carbonmonoxyhemoglobin-lipid interaction has a large electrostatic component. In the glyceraldehyde-3-phosphate dehydrogenase-lipid interaction electrostatic and hydrophobic forces are more equal. The polyhistidine-lipid interaction is almost entirely electrostatic. These results provide information about interactions of cytosolic proteins with lipids.