Exploration Of The Pathogenic Mechanism And Prevention Strategy Of Intracellular Pathogens Based On Chemical Proteomics

The environment we live in is full of various pathogens,which can recognize and invade the host’s body under appropriate conditions,causing infectious diseases and seriously threatening human health.Although the application of antibiotics and interferons can effectively control infections caused by most pathogens,the limitations of this method and the drawbacks of drug resistance have also become increasingly apparent.Therefore,clarifying the pathogenic mechanism and control of pathogens has become one of the core issues in the field of microbiology and immunology,this will also provide a theoretical basis for the development of new drugs and the introduction of appropriate response strategies.In the following paper,we will describe intracellular bacteria and viruses.In Chapter 2,we introduced using chemical proteomics to identify the novel SPI-2 T3 SS effector protein of Salmonella.In this study,we used Salmonella as the chassis cell and inserted the mutant E.coli phenylalanine t RNA synthetase gene into its genome.The mutant phenylalanyl t RNA synthetase with a point mutation of the 294 amino acid position of the wild-type phenylalanine t RNA synthetase,and the amino acid is mutated from the original alanine to glycine(A294G).The mutant alanine t RNA synthetase can specifically recognize unnatural amino acids,and then mark the nascent protein of bacteria.In this study,a stable unnatural amino acid embedding system was constructed to solve the problems of easy loss of transformed plasmids and unstable embedding system encountered in previous studies.In the Salmonella infection host cell model,the goal of labeling the Salmonella nascent proteome in the host cell with unnatural amino acids is achieved;then click chemistryis used to efficiently enrich and enrich the Salmonella nascent protein in the labeled host cell.Highly sensitive identification.Through comparative analysis with the new-born proteome of Salmonella extracellular for one year,and comparison with existing transcriptome data,candidate effector proteins were identified through bioinformatics analysis.This research method realizes the chemical labeling and quantitative analysis of the new proteome of Salmonella in different infection periods in the interaction model of Salmonella infection of host cells,thereby obtaining the spatiotemporal dynamics of the proteome of intracellular pathogens the scientific map reveals the types of effector proteins secreted by Salmonella during different infection periods.The significance of this study lies in the establishment of a chemical proteomic identification method for the novel effector proteins of intracellular pathogens,which lays a foundation for further revealing the infection mechanism of Salmonella and its interaction with host cells,and the interaction between existing pathogens and host cells to supplement the spectra.In Chapter 3,we screened the inhibitors of the main protease of the novel coronavirus and verified that Biotin-VAD-FMK is an active molecular probe for the novel coronavirus.The pandemic of unidentified pneumonia caused by severe acute respiratory syndrome coronavirus 2(SARS-CoV-2)has developed into one of the most serious public health health crises in modern history.SARS-CoV-2 has an impact on global public health and the economy.It has had a very serious impact.So far,the number of patients infected with the virus has exceeded 129 million,and more than2.82 million have died.At present,no specific drug for SARS-CoV-2 has been approved.In order to effectively suppress the further expansion of the epidemic,we have used the main protease,which is essential in the process of virus replication and infection,as the research target.This study combines artificial intelligence models and wet experiments to build a closed loop of mutual feedback,and gradually improves the accuracy of prediction with the model through multiple cycles.The model was used to screen out possible inhibitors from the biologically active chemical library.After two rounds of enzymatic experiments,six new inhibitors and one active molecular probe for SARSCoV-2 Mpro were identified,combined with activity-based the proteomics technology is used to screen active molecular probes and inhibitors of the SARS-CoV-2 main protease,aiming to find compounds that control the new type of pneumonia epidemic and promote the development of anti-coronavirus drugs.