Chemical Biology Study Towards Glycolipid,Gap Junction And Plant Plasma Membrane H~+-ATPase

Studies show increasing evidence that glycolipids are not only important component of cell membrane,but also exert important role in signal transduction.The development of glycolipid probe to unveil their function and transport has attracted the attention of synthetic chemists.In the second Chapter of the thesis,we proposed a novel glycolipid probe design strategy,which features with the substitution of a natural fatty acid by a bifunctional fatty acid equipped with the photoaffinity group diazirine and terminal alkyne.Taking MGDG as an example,we synthesized 3 different MGDG bifunctional probes.And by further acitivity test,we obtained both positive probe and negative probe.By using these probes to do the proteomic profiling study,we revealed,for the first time,that the anti-inflammatory target of MGDG is TLR4.In combination with computational modeling and biochemical experiments,we found that MGDG exert the anti-inflammatory activity in a similar way as known TLR4 antagonist.This probe design strategy can be applied in the study of lipid-protein interaction study.Gap junctions provide a way for direct communication of both energy and information between adjacement cells.The existing methods such as microinjection and FRAP have limitations.The development of a novel fluorescent probe has important significance on the study of gap junctions.In the third chapter of the thesis,we designed a novel fluorocent probe based on the mechanism of?-lactam hydrolysis by?-lactamase.We have obtained 5 different fluorescent probe by chemical synthesis and in vitro studies shows these probes can be hydrolyzed by?-lactamase to release fluorescent.However,the in vivo study showed is not ideal,and further optimization of the probe and the biological system is necessary.Plasma membrane H~+-ATPase is the master enzyme of plants.PM H~+-ATPase can make pH and electric potential to generate energy for the secondary transport system.Because of the gene redundancy,genetic methods have limitiations in the study of PM H~+-ATPase.In the fourth chapter of the thesis,we reported the identification of PM H~+-ATPase inhibitor-PS-1 through high-throughput screening.Further biochemical study revealed that PS-1 can selectively inhibit PM H~+-ATPase.Through structure-activity analysis,we found the modifiable site of PS-1 and synthesized the probe of PS-1equiped with azide group.By using this probe and in combination of other biochemical methods,we found that PS-1 interacts with the centerloop of PM H~+-ATPase to exert its inhibition activity.PS-1 is an important chemical tool to study the function of PM H~+-ATPase...