Structural And Functional Studies Of 3-ketodihydrosphingosine Reductase In Ceramide Metabolism

Ceramide(Cer)is an important active lipid that regulates cell proliferation,differentiation,apoptosis and other major life activities,and is the center of sphingolipid metabolism.Because ceramide plays an important role as a second messenger in cell signaling pathways,its synthesis and degradation in the body are strictly regulated,and its content is in a dynamic balance,its synthesis includes de novo synthesis pathways,sphingomyelinase pathways,and salvage pathways.In recent years,studies have shown that enzymes in the de novo synthesis pathway are associated with many diseases such as obesity,rare diseases,and cancer,which illustrates the complexity and importance of this pathway in the body's life activities.3-Ketodihydrosphingosine reductase(KDSR)is a three-transmembrane protein localized on the endoplasmic reticulum and is responsible for the reduction of 3-ketodihydrosphingosine to dihydrosphingosine in the de novo synthesis pathway of ceramide.Many studies have shown that KDSR mutation will cause a rare skin disease,namely progressive symmetric erythrokeratodermia(PSEK),and may be accompanied by other diseases such as thrombocytopenia.However,the catalytic mechanism and pathogenic mechanism of KDSR are currently unclear.In this regard,this article takes KDSR as the experimental object,using molecular biology,structural biology,biochemistry and other technical methods,first expressed and purified wild-type KDSR and three pathological mutants(p.F138 C,p.A175 T,p.Y186F)with E.coli prokaryotic expression system,and then analyzed the high-resolution three-dimensional structures of KDSR and three pathological mutants,indicating local differences in the structure of wild-type and mutants.At the same time,the thesis also established an enzyme activity measurement system to characterize the in vitro enzyme activity of wild-type and mutants.The above studies have initially clarified the pathological pattern of KDSR,which has laid the foundation for the in-depth study of the pathogenesis of KDSR,and also has important guiding value for the treatment of the rare disease PSEK.