On The Interactions Of Human Apolipoprotein H With Phospholipid Membranes And The Conformation Of Membrane-bound Apolipoprotein H

Apolipoprotein H (ApoH) is a plasma glycoprotein belonging to the family of apolipoprotein. Hitherto, the biological function of ApoH is not very clear. Since it is known that the in vivo physiological and pathogenic roles of ApoH are closely related to its interaction with the phospholipid membrane, a more profound understanding on the interaction mechanism, as well as on the membrane-bound conformation of ApoH may have not only theoretically but also practically beneficial consequence. In the dissertation, the interaction of ApoH with phospholipid membranes and the conformation of membrane-bound ApoH was studied by using different model systems of lipid monolayer, liposome and an organic solvent contain water and methanol. Firstly, with the method of phospholipid monolayer at air/water interface, we proved for the first time that ApoH is an amphiphilic protein which is capable of insertion into negatively charged lipid monolayer. The membrane insertion depth of ApoH is about 2± A. In the process of ApoH insertion into membranes, both the electrostatic and the hydrophobic interactions between protein and the lipid are evidently involved. Secondly, we combined techniques of fluorescence and circular dichroism spectroscopy to study the conformation of membrane-bound ApoH. The experimental results showed that after binding to the negatively charged liposome, the secondary structure of ApoH will be significantly changed and some a-helix structure can be induced. By changing the composition of phospholipid in vesicles and the ionic strengths in the buffer, we found that the membrane-bound ApoH molecules can adopt two conformations at the membrane surface. Under conditions of low ionic strength, the water-soluble ApoH will bind to the negatively charged lipid vesicles with most of its tryptophan residues adsorbing on the membrane surface and a small part of them inserting into the membrane. Under conditions of high ionic strength, the membrane-bound ApoH will have a different conformation with its non-inserted tryptophan residues mainly separated from the membrane surface, and its membrane- inserted ones still maintained their association with the membrane. The conformational change of ApoH upon binding to the membrane may provide important structural bases for the protein's biological function, e.g. in clearing the negatively charged liposome in plasma, and in the autoimmune diseases such as systemic lupus erythematosus. Thirdly, we studied the correlation between the membrane insertion ability and the membrane-bound conformation of ApoH with the model system of water- methanol mixed solvent at moderately low pH that mimics the micro- physicochemical environment near the membrane surface. In a buffer of 57% methanol, ApoI-I can be induced to adopt a special conformation just the same as that bound to the negatively charged membrane. On the other hand, it is observed that the membrane insertion ability of the conformational changed ApoH increased, too. With the method of capillary electrophoresis, it is found that in this case, there are more ApoH molecules adsorbing on the monolayer surface. The above experiments suggest the close relationship between the membrane insertion ability and the membrane- bound conformation of ApoH. In all, we partially revealed the molecular mechanism of the interaction of ApoH with phospholipid membranes and obtained a model for the membrane insertion and conformational change of ApoH at the membrane surfa...