Screening And Preliminary Verification Of Affinity Peptide Of BMP2 Receptor Protein

Bombyx mori silk fibroin is an excellent biomaterial,which has been widely concerned and researched for its degradability and good biocompatibility,and its application range is also wider and wider.However,the implementation of numerous applications of silk fibroin is dependent on the coupling of various functional factors and other materials.The current coupling method is primarily a simple physical mixing or chemical coupling.Simple physical mixing has the problem of uneven mixing and weak combination.The chemical coupling method is complicated,inefficient,and cannot be detached after connection.To solve this problem,this thesis uses the phage peptide library screening technology to find the specific binding peptide of silk fibroin,and finds one of the strongest binding ability through a series of verification experiments.This peptide is expected to be used as a bridge peptide to link other materials to silk fibroin and broaden its application in the field of biomaterials.Bone morphogenetic protein 2(BMP2)is a functional factor that promotes the differentiation of bone marrow mesenchymal stem cells into osteoblasts.But,it is expensive on the market and has limited sources.This has made it impossible to be widely used.To solve this problem,this thesis uses the phage peptide library screening technology to find the peptide specifically recognized by its receptor protein,and displays the peptide with the best affinity on the phage p? coat protein.The phage was further prepared into a topological membrane to explore the effect on stem cells.The main works of this paper are as follows:(1)A peptide specifically binding to silk fibroin was obtained by affinity screening experiments of phage random 12 peptide library and silk fibroin.The HVAWSWSWNNST peptide sequence with the strongest binding was identified by phage capture assay and phage-enzyme-linked immunosorbent assay.(2)Affinity peptides of two receptor proteins were obtained by affinity screening experiments of C7C phage random peptide library and two receptor proteins BMPR2 and BMPR1A of BMP2,respectively.The phage capture assay and phage-enzyme-linked immunosorbent assay showed that the affinity of SLFSKN peptide and BMPR2 protein was the best,and the affinity of KRKYRRG peptide and BMPR1A protein was the best.(3)Engineered phage displaying the BMP2 receptor protein affinity peptide were obtained by phage display technology and recombinant plasmid transfection.A nanoscale phage membrane with unique ridge-groove morphology was prepared by the pulling method.The optimal phage concentration was 1013 pfu/ml and the optimal pulling speed was 50 ?m/min.The phage display peptide had no effect on the membrane morphology.Different pulling bases have an effect on the membrane morphology.When the membrane surface was coated with poly-lysing,the phage morphology structure can be effectively immobilize,so it can be stably present in a solution environment.(4)The phage membrane has good biocompatibility by cell culture experiments.