家蚕丝素蛋白亲和多肽的筛选及验证
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摘要
家蚕丝素蛋白因其良好的生物相容性被广泛以修饰或基底材料的形式用于生物医用复合材料或器件,但与其他材料的结合性和相容性有待提高。通过噬菌体多肽库淘选技术筛选出家蚕丝素蛋白亲和肽有望解决这个问题。用噬菌体随机多肽库对家蚕丝素蛋白进行亲和筛选,获得多个家蚕丝素蛋白亲和多肽,其中亲和性最好的多肽序列为HVAWSWSWNNST,其出现的频率为42%。通过生物信息学软件RELIC/info验证了筛选过程的有效性。HVAWSWSWNNST多肽组在噬菌体捕获实验中的噬菌体回收数量最多且在噬菌体-酶联免疫吸附实验中具有最高的吸光度值,证明该多肽与丝素蛋白的结合力最强。该多肽的成功筛选,有望改善家蚕丝素蛋白与其他材料的结合性和相容性,并拓宽丝素蛋白在生物医学领域的应用。
Silkworm fibroin is widely used in biomedical composite materials or devices as a modified or substrate material due to its good biocompatibility. But its binding ability and compatibility with other materials need to be improved. Affinity peptide screening for silkworm fibroin by phage peptide library panning technology is expected to solve this problem.In this study,a few affinity peptides for silkworm fibroin were obtained by affinity screening in phage random peptide library,among which the peptide HVAWSWSWNNST showed the best affinity with a frequency of 42%. Effectiveness of this screening process was verified by bioinformatic analysis. Since HVAWSWSWNNST peptide group had the biggest amount of recovered phages in the phage capture experiment,and the highest absorbance in phage-enzyme linked immunosorbent assays,it was suggested that its binding ability to silkworm fibroin was the strongest. The screened peptide is expected to improve the binding ability and compatibility of silk fibroin with other materials,and broaden its application in biomedical fields.
Silkworm fibroin is widely used in biomedical composite materials or devices as a modified or substrate material due to its good biocompatibility. But its binding ability and compatibility with other materials need to be improved. Affinity peptide screening for silkworm fibroin by phage peptide library panning technology is expected to solve this problem.In this study,a few affinity peptides for silkworm fibroin were obtained by affinity screening in phage random peptide library,among which the peptide HVAWSWSWNNST showed the best affinity with a frequency of 42%. Effectiveness of this screening process was verified by bioinformatic analysis. Since HVAWSWSWNNST peptide group had the biggest amount of recovered phages in the phage capture experiment,and the highest absorbance in phage-enzyme linked immunosorbent assays,it was suggested that its binding ability to silkworm fibroin was the strongest. The screened peptide is expected to improve the binding ability and compatibility of silk fibroin with other materials,and broaden its application in biomedical fields.
引文
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[2]BABA A,KITAYAMA K,ASAKURA T,et al.Silk fibroin produced by transgenic silkworms overexpressing the Arg-Gly-Asp motif accelerates cutaneous wound healing in mice[J/OL].J Biomed Mater Res,B,2016,84(1):e150[2019-01-15].https://www.ncbi.nlm.nih.gov/pubmed/29504231.DOI:10.1002/jbm.b.34098
[3]YI B,ZHANG H,YU Z,et al.Fabrication of high performance silk fibroin fibers via stable jet electrospinning for potential use in anisotropic tissue regeneration[J].J Mater Chem,B,2018,6:3934-3935
[4]MONTALBAN M G,COBURN J M,LOZANO-PEREZ A A,et al.Production of curcumin-loaded silk fibroin nanoparticles for cancer therapy[J/OL].Nanomaterials,2018,8(2):126[2019-01-15].https://www.ncbi.nlm.nih.gov/pubmed/29495296.DOI:10.3390/nano8020126
[5]LI X T,XIE H Q,CHEN Y J,et al.Silkworm pupa protein hydrolysate induces mitochondria-dependent apoptosis and S phase cell cycle arrest in human gastric cancer SGC-7901 cells[J/OL].Int JMol Sci,2018,19(4):e1013[2019-01-15].https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5979490/.DOI:10.3390/ijms190-41013
[6]LING S J,LI C X,ADAMCIK J,et al.Directed growth of silk nanofibrils on graphene and their hybrid nanocomposites[J].ACSMacro Lett,2014,3(2):146-152
[7]NILEBACK L,CHOUHAN D,JANSSON R,et al.Silk-silk interactions between silkworm fibroin and recombinant spider silk fusion proteins enable construction of bioactive materials[J].ACS Appl Mater Interfaces,2017,9(37):31634-31644
[8]HU K,GUPTA M K,KULKARNI D D,et al.Ultra-robust graphene oxide-silk fibroin nanocomposite membranes[J].Adv Mater,2013,25(16):2301-2307
[9]ARMSTRONG J,PERHAM R N,WALKER J E.Domain structure of bacteriophage fd adsorption protein[J].FEBS Lett,1981,135(1):167-172
[10]DEMARTIS A,LAHM A,TOMEI L,et al.Polypharmacy through phage display:selection of glucagon and GLP-1 receptor coagonists from a phage-displayed peptide library[J/OL].Sci Rep,2018,8:585[2019-01-15].https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5766609/.DOI:10.1038/s41598-017-18494-5
[11]DE PlANO L M,CARNAZZA S,MESSINA G M L,et al.Specific and selective probes forStaphylococcus aureusfrom phagedisplayed random peptide libraries[J].Colloids Surf,B,2017,157:473-480
[12]YOU F,YIN G,PU X,et al.Biopanning and characterization of peptides with Fe3O4nanoparticles-binding capability via phage display random peptide library technique[J].Colloids Surf,B,2016,141:537-545
[13]LI Y,CAO B R,YANG M Y,et al.Identification of novel short BaTiO3-binding/nucleating peptides for phage-templated in situ synthesis of BaTiO3polycrystalline nanowires at room temperature[J].ACS Appl Mater Interfaces,2016,8(45):30714-30721
[14]LI J,FENG L,FAN L,et al.Targeting the brain with PEG-PLGAnanoparticles modified with phage-displayed peptides[J].Biomaterials,2011,32(21):4943-4950
[15]HOU L,ZHU D,LIANG Y,et al.Identification of a specific peptide binding to colon cancer cells from a phage-displayed peptide library[J].Br J Cancer,2017,118(1):79-87
[16]WANG S,HUMPHREYS E S,CHUNG S,et al.Peptides with selective affinity for carbon nanotubes[J].Nat Mater,2003,2(3):196-200
[17]KASE D,KULP J L,YUDASAKA M,et al.Affinity selection of peptide phage libraries against single-wall carbon nanohorns identifies a peptide aptamer with conformational variability[J].Langmuir,2004,20(20):8939-8941
[18]NAIK R R,STRINGER S J,AGARWAL G,et al.Biomimetic synthesis and patterning of silver nanoparticles[J].Nat Mater,2002,1(3):169-172
[19]SARIKAYA M,TAMERLER C,JEN A K Y,et al.Molecular biomimetics:nanotechnology through biology[J].Nat Mater,2003,2(9):577-585
[20]MAO C,FLYNN C E,HAYHURST A,et al.Viral assembly of oriented quantum dot nanowires[J].Proc Natl Acad Sci USA,2003,100(2):6946-6951
[21]MAO C,SOLIS D J,REISS B D,et al.Virus-based toolkit for the directed synthesis of magnetic and semiconducting nanowires[J].Science,2004,303(5655):213-217
[22]MANDAVA S,MAKOWSKI L,DEVARAPALLI S,et al.RELIC-a bioinformatics server for combinatorial peptide analysis and identification of protein-ligand interaction sites[J].Proteomics,2004,4(5):1439-1460
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