Identification Of Enzymes Involved In An Anaerobic Purine Degradation Pathway And Research On Multifunctional Nanocarriers Based On MoS₂ For Cancer Therapy

Purines are nitrogen-containing heterocyclic aromatic compounds that are important components of nucleic acids and many crucial metabolic cofactors,which makes them essential for all life.Purines play important roles in the cellular energy supply,signal transduction and enzyme catalysis.When animals ingest excess purines,they must be metabolized and excreted.Primates are different from other mammals in purine metabolism because they lack the uricase gene that enables uric acid degradation,so the end-product of purine metabolism in primates,including humans,is uric acid.A high content of uric acid in the blood can cause it to crystallize in the joints,which in turn induces an inflammatory reaction and causes gout.Microorganisms can use purines as a carbon and nitrogen source,as well as obtain energy through their complete mineralization.All known purine degradation pathways transform xanthine to uric acid,which is catalyzed by xanthine oxidase or xanthine dehydrogenase containing molybdenum and sulfur complexes as auxiliary groups,with either O₂ or NAD+as electron acceptor,respectively.Through the method of bioinformatics,we found that the gene cluster of purine transporter gene encoded by Bacillus firmus contains new enzymes and metabolic pathways that may catalyze purine degradation.On this basis,I cloned,expressed and purified two enzymes of this pathway,and through the enzyme activity experiment in vitro,we detected that xanthine was catalyzed by the amide hydrolase family to hydrolyze the ring to produce 4-urea-5-carboxyimidazole.the latter further through the action of the second enzyme to produce4-amino-5-carboxyimidazole,thus analyzing the mechanism of the previous detection of such intermediate metabolites in cell metabolites.This work is of great significance in theory:the purine degradation pathway independent of xanthine oxidase/dehydrogenase was discovered for the first time in nature.At the same time,it has important potential application value:the new enzyme does not need oxygen and complex auxiliary groups,and can be overexpressed and active in intestinal bacteria.Through genetic engineering of intestinal probiotics,excess purines in food can be degraded in advance in the intestine,so as to reduce the content of blood uric acid,treat and prevent gout.With the rise of nanotechnology,the application of nano-medicine in the treatment of tumors has attracted wide attention,but there are still many factors restricting its development in the imaging-guided treatment of tumors.For example,it is difficult to completely eliminate the tumor with a single treatment,and accurate imaging technology is needed to judge the location,size and distribution of the tumor.Additionally,some nanoparticles cannot be completely removed from the body after treatment.In this study,a multifunctional nanoparticle was constructed,which can passively target the tumor by enhancing the penetration and retention effect,realize bimodal tumor therapy guided by multimode imaging technology,and catalyze redox reactions with the highly concentrated GSH in the tumor microenvironment,resulting in structural rupture of the nanoparticles and final clearance.