STR1Based Expression System Optimization,Substrate Specificity Study And Chemoenzymatic Approach To Novel Indole Alkaloids&Heterocylic Compounds Construction

Strictosidine synthase (STR) is one of the most important enzymes for monoterpenoid indole alkaloids (MIAs) biosynthesis, which catalyzes the stereoselective Pictet-Spengler reaction of tryptamine and secologanin to form the key intermediate3α(S)-strictosidine in MIAs biosynthesis. MIAs, derived from strictosidine, have intensive biological activities and some of them have been in clinical use. Due to the multi-chiral centers and polycyclic rings they have, it is always challenging to synthesize them by traditional synthetic approaches. Currently, plants are the main source of those drugs coming from MIAs.The second part of the thesis has a description of STR1based expression system optimization, substrate specificity study and chemoenzymatic approach to novel indole alkaloids. In order to get novel terpenoid indole alkaloids molecular diversity and potential biological activities, pretty much amount of strcitosidine and its derivative are in demand which will rely on improved expression system of STR1. An improved systematic research on STRl expression with chaperones was carried out and chaperone pG-Tf2contributed to the doubled expression of soluble His6-STR1expression in Escherichia coli. Meanwhile, series of tryptamine analogues were designed and synthesized to check the substrate specificity of STRl in which some of them can be accepted by STR1. We also described the mechanism of STR1catalyzed transformation of unique substrate to new strictosidine using3D crystal structures of STRl in a complex with the substrate or product. After that, the immobilized STR1on Ni-NTA was used to prepare strictosidine and its novel analogues. These strictosidines can be used as raw materials to afford novel indole alkaloids by "One-pot" biomimetic synthesis with the existence of acid, base, β-glucosidase and NaBH4respectively. Some TopoI inhibitors based on these alkaloids were designed and partially synthesized.Heterocycles usually have a wide range of biological activities. In the third part of the thesis, we developed the methodologies to afford several heterocyclic compounds, including a facile approach to provide polysubstituted pyrazoles,"One-pot" multicomponent way to obtain indolizines and pyrido[1,2-a]indoles as well as construction of five heterocycles from Morita-Baylis-Hillman Acetates. Through random screening, we found that some indolizines have better inhibition on TRPM2, the important target of apoptosis of dopamine neurons induced by reactive oxygen species (ROS), than those reported small molecues. These indolinzines might be used as small molecule probes to study the central nervous system disease and neurodegenerative disease.