Bioinformatics Research And Application Of Phage Display

Phage display is a high-throughput screening technology based on in vitro display,which allows for the efficient enrichment of peptides or proteins with specific functions through iterative affinity selection(biopanning)from libraries with billions of peptides.In recent years,next-generation sequencing(NGS)has been combined with phage display to form next-generation phage display(NGPD)that has greatly contributed to the analysis of biopanning results.Phage display has been widely used in basic and applied research,such as epitope prediction,protein interaction network analysis,and the development of disease diagnosis and treatment products,drug delivery agents,novel biological and inorganic functional materials.The quality of the phage display library is crucial for the success of biopanning.With the development of sequencing technology,NGS analysis allows for the evaluation of the quality of libraries with 105-106 diversity.However,NGS technologies are mainly used for genome sequencing data analysis.Most of the available software is written for genome assembly.There is a lack of software tailored for analyzing phage display libraries and evaluating the quality of the large-scale libraries.Therefore,there is an urgent need in the field of phage display to develop analytical procedures for assessing the library quality.Due to the internal causes of the phage display panning system and library,phage display selection results contain target-unrelated peptides(TUPs)as well as target-specific binding peptides.Researchers in the community have difficulty in distinguishing these two types of peptides.Computational methods and tools have the advantages of high efficiency and low cost.Therefore,the identification of noisy sequences by bioinformatics tools has played an important part in the field of phage display.To respond to the need and development of the field,a new paired-end processing program that is special for the analysis of phage display libraries is developed.A standard analytical platform for assessing and characterizing the diversity of phage-displayed libraries is also proposed.To eliminate TUPs from phage display results,a series of computational tools to diagnose and report TUPs is developed and improved.In addition,the interaction between metuximab and CD 147 is investigated using phage display and bioinformatics tools.The main research contents of the thesis are as follows:Firstly,a paired-end processing platform and an analytical workflow for evaluating the library diversity are proposed.The newly developed paired-end processing program is employed to study the compositional bias in naive and chemically modified phage-displayed libraries.Secondly,TUP reporting tools based on the biopanning database are improved.Besides the traditional phage display data,large-scale next-generation phage display data are also curated.A series of TUP analysis tools is integrated into the Biopanning Data Bank(BDB).The website of the database is also redeveloped and improved to be more user-friendly.BDB provides an evidence-based platform to help users identify TUPs within phage display data.Thirdly,TUP prediction tools based on machine learning methods,namely SABinder and PhD7Faster 2.0,are developed.Streptavidin-binding peptides(SBP)are collected from the BDB database,and a tool called SABinder is developed based on Support Vector Machine(SVM)to predict SBP.PhD7Faster is also improved and upgraded to version 2.0 to predict propagation-related TUPs in the PhD-7 phage display library.SABinder and PhD7Faster 2.0 have been developed into web applications that provide free online predicting services.In addition,a standalone version has also been developed and provided to facilitate the predictive analysis of large datasets.Fourthly,the TUP analysis tool suite,SAROTUP,is redeveloped.Many new TUP motif sequences are integrated into the TUPScan tool and three SVM-based tools,SABinder,PhD7Faster 2.0 and PSBinder,are integrated into the latest version of SAROTUP.We have also developed a graphical user interface and command-line version of the SAROTUP application for large-scale biopanning data analysis and released it as an open source package.The website of SAROTUP has been redesigned.Users can freely access SAROTUP through http://i.uestc.edu.cn/sarotup3.Fifthly,a study case is performed using phage display.Metuximab is used to screen the PhD-12 phage display library,and its epitope is analyzed by homology modeling,molecular docking and interpreting data from phage-displayed library screening.In summary,the thesis focuses on bioinformatics research on phage display.A series of computational tools for phage display data analysis is proposed.Finally,phage display and bioinformatics tools are employed to study the interaction between metuximab and CD147.The bioinformatics resource and tools developed in this project will accelerate the development of new drugs,vaccines,diagnostic reagents and materials based on phage display,which have important scientific significance and application prospects.