| Caveolae which is about 50-100nm various invaginations exists on the surface of cell plasma membrane.Most signal transduction protein enriched on that and responsible for many biological functions.It is the center of cell signal processing.Mainly carring the signal transduction inhibitor. Caveolin are Caveolae's characteristic protein, which can fix signal molecules by forming scaffold when the cell in a steady state or the signal behavior leading by ligand. As the permanent structure on the plasma membrane, the research on physiological function of caveolae and Caveolin are getting more and more attention. especially in signal transduction,cholesterol transport,tumor suppression,muscle cell synthesis and so on.Caveolae play an important role in the process of cell activities and which have great research value.In this paper,we have researched Caveolae and lipis interactions in two respects: (1)The separation and identification of Caveolae. (2) Preliminary study on simulation of artificial Plasma membrane in vitro. Through to the two aspects of research work which could lay a good foundation for the further researchon the structure and functions of caveolae,and have great significance.The experiment results are as follows:(1) The separation and identification of Caveolae:We separated caveolae from fresh pig lungs by using the nonionic detergents TritonX-100 which caveolae can not dissolve in it at 4℃. SDS-PAGE rusults the line of the caveolae protein,then we tested caveolin-1 singnal by spothybridization.Through the test of Coomassie Brilliant Blue, we get the protein content is 0.79113mg/ml. Then we observed them by atomic force microscopy (AFM) in different conditions and the best AFM images of caveolae were obtained by diluting them 500 times with deionized water.(2) Preliminary study on simulation of artificial Plasma membrane in vitro:In order to study the mechanism of the interaction between different kinds of lipid which constituted caveolae,we simulated plasma membrane in vitro. The physical properties and morphology of mixed binary systems of phosphatidylcholine dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) with Cholesterol (Chol) have been investigated at the air-water interface by surface pressure-area (π-A) isotherm measurements and by atomforce microscopy(AFM) observation. The analyses of theπ-A isotherms show that at 20(±0.5)℃the areas of the binary systems decrease with the increasing surface pressure (π) when the DPPC molar fraction(XDPPC) keeps invariant;when the surface pressure keeps invariant,at the surface pressure of 15mN/m, the areas of the binary systems increase at first,and then decrease. but at the surface pressure of 20mN/m,25mN/m,30mN/m, the areas of the binary systems decrease nonlinearly and periodically with the increasing DPPC molar fraction. The analysis of the Gibbs energy indicate that the systems obtain maximum negative deviation at XDPPC=0.7; but at the surface pressure of 20mN/m,25mN/m,30mN/m, the systems obtain conspicuously maximum positive at XDPPC=0.1.These results have shown the characteristic of XDPPC vary withπ,and have been directly confirmed by AFM observation. Finally,We chose the different proportion of the binary systems, and loaded the caveolae onto it.Then compare the difference between loaded and unloaded. Analysis the way of interaction between caveolae and monolayer,and observed the whole process through the AFM and know about the distribution and presentation of the caveolae.This study has important biological significance for further investigations of the liquid crystal state variations of biomembrane.This part studied on the phase behavior and morphology of mixed binary systems of DPPC/Chol at the air/water interface by experiment researches and theoretical researches,so this research had important meaning of exploration.This paper provided important experimental basis and new idea for the further research on the structure and functions of caveolae,and had great significance. |
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