In Vitro Reconstitution And Optimization Of The Entire Pathway To Convert Glucose Into Fatty Acid

Glucose and fatty acids play crucial roles in almost all living organisms.Glucose is such an important and common energy sources that used by almost all organisms.Glycolysis is an efficient way that used by the body under the anaerobic conditoion.Fatty acids are crucial components of cells as they play important roles as structural components of membranes,energy storage molecules,intracellular messengers,precursors of hormones and protein post-translational modifications moieties.Besides,glucose and fatty acids are also have close relationships with the genesis and development of metabolic disorders such as obesity,diabetes mellitus and cardiovascular diseases.The pathway from glucose to fatty acid covers the most basic and important metabolic patheway of living organisms,which make it extrimely important to profoundly and comprehensively understand the entire pathway and clarify the metabolic mechanism of glucose and fatty acids.So our effort will help humans better understand the genesis and development of metabolic diseases.In our study,we successfully reconstituted in vitro the entire pathway that converts glucose to fatty acid for the first time.This whole pathway consisting of 30 enzymes has 19-step reactions and can be divided into three major systems.In order to improve the efficiency of the whole system,we systematicly optimized the glycolytic pathway and pyruvate dehydrogenase.In order to improve the supply of NADPH in our system,we reconstitutioned in vitro the entire pentose-phosphate pathway of Escherichia coli.The reconstitution of pentose phosphate-glycolysis hybrid pathway and the introduction of GAPDH of Clostridiumacetobutylicum not only improved the supply of NADPH,but also enabled the stability of the whole system,at the same time verified the potential of cofactor recycling of reconstitution system in vitro.At last we increased the yield of free fatty acid from non-detectable by GC-MS to 9.3%of the theoretical yield.From the information we got from the reconstituted in vitro system that converts glucose into fatty acids,we are able to take a more profoundly and comprehensively insight of the metabolism mechanism,and deepen our understanding of the whole complex biological metabolic system.With the apparent advantage of flexible-operating,easy-controlling,our system can not only provide an excellent platform for fundemental biochemical research,but also present a new strategy in drug screening of compounds related to these essential central pathways.Wrapped into liposome which acts as a carrier of the drug-delivery system,our system can promote energy supply and self repairment,promoting the idea of oral delivering protein drug like insulin become a reality.