Study On Extreme PH,Poly-L-Lysine Hydrobromide And Thermal Response Of Phospholipid Bilayers

Biomembranes,composed of proteins and phospholipid bilayers,separate the cells/organelles and their outside environment.They participate in material internalization,energy transfer,signal transduction and many other important life activities.And a comprehensive understanding of how the membranes are composed,structured,functioning and interacting with the life materials,is critical to the applications of the membranes in biomedicine,industry,and agriculture areas.Artificial lipid bilayers are simple biomembrane model employed in membrane-related function and structure researches.In this thesis,we have studied the effects of electric field,polyelectrolyte,medium(pH and temperature)on the structure and assembly of lipid membranes with the inverted fluorescence microscopy,quartz crystal microbalance and dissipation monitor(QCM-D),atomic force microscopy,and dynamic light scattering methods.In the first chapter,we reviewed the basic understandings in the biomembranes and model lipid membranes,such as their compositions,structures,properties,and functions.Then we focused on two common model membranes,supported lipid bilayers(SLBs)and giant unilamellar vesicles(GUVs),introducing the recent research progresses on one important poly-L-Lysine hydrobromide(PLL)and membrane interactions based on the platforms.In the second chapter,we summarized the primary characterization techniques used in this thesis,including the fluorescence microscopy,QCM-D,home-made flow chamber,and particularly the electrofonnation method of GUVs.Specifically,we employed the fluorescence microscopy and home-made flow chamber to monitor the electroformation of GUVs in real time.The GUVs formation properties were found to be decided by the electric field,homogeneity of the dried lipid layers,types of lipids,buffers,and temperature.Due to the polarity of lipid head groups,electric field induces the asymetry of the lipid leaflets,and thus bending,budding.In the third chapter,we studied the effects of exposing GUVs to PLL.We found PLL induced the formation of bright rod-like structures in the interior of the negatively charged GUVs and the binding,exocytosis,and rupture of membranes.In pH 6.98,PLL did not cause shape changes in neutralized charged GUVs.The interactions of PLL and negatively charged GUVs were affected by the concentration distribution of PLL.In constant PLL concentration,the alkaline environment(experiments in pH = 5.60,6.98,8.05)was favored for the interactions,which induced fast binding and rupture of membranes even at an extremely low PLL concentration.The conglutination of GUVs forming in pH = 5.60 can gradually be improved by the addition of PLL.But rod-like structures and rupture were not observed at this pH.We attributed electrical interaction between lipid head groups and PLLs to the responses of GUVs to PLL.In the fourth chapter,we employed fluorescence microscopy to study the SLB assembly caused by the pH shift from the neutral conditions to strong acidity(or alkalinity).The extreme acidity inducing rupture and alkalinity inducing platform,tubulation of membranes were interpreted.From analyzing the factors critical to membrane shape change,we proposed that the mechanism undying the assembly dynamics of lipid bilayer under extremely pH environments.In the fifth chapter,we studied the membrane assembly when thermal condition changed.Firstly,the SLB mass and lateral diffusivity were found to impair by a heating process.3-D fluorescence reconstitution showed that membrane fission is the origin of the membrane integrity disruption,which is generated by the heating induced lipid lateral area expansion.Next,the salt concentration in the medium was found to be able to regulate the heating-related-SLB-disruption.The high ionic concentration inhibits the spontaneous release of membrane tension and membrane fission.Theory analysis displays that the osmotic energy dominates the fission process.The semi-permeability of lipid bilayers results in the osmotic imbalance during membrane budding,which boosts with the increase of the bulk solute concentration.Finally,the osmotic-based SLB-heating-protection strategy was demonstrated to be solute independent,side-effect low.