Angelika Becht, Justyna Frączyk, Joanna Waśko, Elżbieta Menaszek, Jakub Kajdanek, Katarzyna Miłowska, Beata Kolesinska
{"title":"挑选形成原生蛋白外层的胶原蛋白 IV 片段:评估再生医学的生物活性。","authors":"Angelika Becht, Justyna Frączyk, Joanna Waśko, Elżbieta Menaszek, Jakub Kajdanek, Katarzyna Miłowska, Beata Kolesinska","doi":"10.1002/psc.3537","DOIUrl":null,"url":null,"abstract":"<p>The aim of this research was to select the fragments that make up the outer layer of the collagen IV (COL4A6) protein and to assess their potential usefulness for regenerative medicine. It was expected that because protein–protein interactions take place via contact between external domains, the set of peptides forming the outer sphere of collagen IV will determine its interaction with other proteins. Cellulose-immobilized protein fragment libraries treated with polyclonal anti-collagen IV antibodies were used to select the peptides forming the outer sphere of collagen IV. In the first test, 33 peptides that strongly interacted with the polyclonal anti-collagen IV antibodies were selected from a library of non-overlapping fragments of collagen IV. The selected fragments of collagen IV (cleaved from the cellulose matrix) were tested for their cytotoxicity, their effects on cell viability and proliferation, and their impact on the formation of reactive oxygen species (ROS). The studies used RAW 264.7 mouse macrophage cells and Hs 680.Tr human fibroblasts. PrestoBlue, ToxiLight™, and ToxiLight 100% Lysis Control assays were conducted. The viability of fibroblasts cultured with the addition of increasing concentrations of the peptide mix did not show statistically significant differences from the control. Fragments 161–170, 221–230, 721–730, 1331–1340, 1521–1530, and 1661–1670 of COL4A6 were examined for cytotoxicity against BJ normal human foreskin fibroblasts. None of the collagen fragments were found to be cytotoxic. Further research is underway on the potential uses of collagen IV fragments in regenerative medicine.</p>","PeriodicalId":16946,"journal":{"name":"Journal of Peptide Science","volume":null,"pages":null},"PeriodicalIF":1.8000,"publicationDate":"2023-08-22","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":"{\"title\":\"Selection of collagen IV fragments forming the outer sphere of the native protein: Assessment of biological activity for regenerative medicine\",\"authors\":\"Angelika Becht, Justyna Frączyk, Joanna Waśko, Elżbieta Menaszek, Jakub Kajdanek, Katarzyna Miłowska, Beata Kolesinska\",\"doi\":\"10.1002/psc.3537\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<p>The aim of this research was to select the fragments that make up the outer layer of the collagen IV (COL4A6) protein and to assess their potential usefulness for regenerative medicine. It was expected that because protein–protein interactions take place via contact between external domains, the set of peptides forming the outer sphere of collagen IV will determine its interaction with other proteins. Cellulose-immobilized protein fragment libraries treated with polyclonal anti-collagen IV antibodies were used to select the peptides forming the outer sphere of collagen IV. In the first test, 33 peptides that strongly interacted with the polyclonal anti-collagen IV antibodies were selected from a library of non-overlapping fragments of collagen IV. The selected fragments of collagen IV (cleaved from the cellulose matrix) were tested for their cytotoxicity, their effects on cell viability and proliferation, and their impact on the formation of reactive oxygen species (ROS). The studies used RAW 264.7 mouse macrophage cells and Hs 680.Tr human fibroblasts. PrestoBlue, ToxiLight™, and ToxiLight 100% Lysis Control assays were conducted. The viability of fibroblasts cultured with the addition of increasing concentrations of the peptide mix did not show statistically significant differences from the control. Fragments 161–170, 221–230, 721–730, 1331–1340, 1521–1530, and 1661–1670 of COL4A6 were examined for cytotoxicity against BJ normal human foreskin fibroblasts. None of the collagen fragments were found to be cytotoxic. Further research is underway on the potential uses of collagen IV fragments in regenerative medicine.</p>\",\"PeriodicalId\":16946,\"journal\":{\"name\":\"Journal of Peptide Science\",\"volume\":null,\"pages\":null},\"PeriodicalIF\":1.8000,\"publicationDate\":\"2023-08-22\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"\",\"citationCount\":\"0\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"Journal of Peptide Science\",\"FirstCategoryId\":\"99\",\"ListUrlMain\":\"https://onlinelibrary.wiley.com/doi/10.1002/psc.3537\",\"RegionNum\":4,\"RegionCategory\":\"生物学\",\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"\",\"PubModel\":\"\",\"JCR\":\"Q4\",\"JCRName\":\"BIOCHEMISTRY & MOLECULAR BIOLOGY\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"Journal of Peptide Science","FirstCategoryId":"99","ListUrlMain":"https://onlinelibrary.wiley.com/doi/10.1002/psc.3537","RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q4","JCRName":"BIOCHEMISTRY & MOLECULAR BIOLOGY","Score":null,"Total":0}
引用次数: 0
摘要
这项研究的目的是选择构成胶原蛋白 IV (COL4A6) 蛋白外层的片段,并评估它们在再生医学中的潜在用途。由于蛋白质与蛋白质之间的相互作用是通过外部结构域之间的接触进行的,因此构成胶原蛋白 IV 外层的一组肽将决定其与其他蛋白质的相互作用。使用多克隆抗胶原蛋白 IV 抗体处理的纤维素固定化蛋白质片段库用于筛选形成胶原蛋白 IV 外球的肽。在第一次试验中,从胶原蛋白 IV 的非重叠片段库中筛选出 33 个与多克隆抗胶原蛋白 IV 抗体有强烈相互作用的肽段。对所选的胶原蛋白 IV 片段(从纤维素基质中裂解)进行了细胞毒性、对细胞活力和增殖的影响以及对活性氧(ROS)形成的影响测试。研究使用了 RAW 264.7 小鼠巨噬细胞和 Hs 680.Tr 人成纤维细胞。进行了 PrestoBlue、ToxiLight™ 和 ToxiLight 100% 裂解对照试验。加入浓度不断增加的多肽混合物培养的成纤维细胞的存活率与对照组相比没有显著的统计学差异。检测了 COL4A6 的 161-170、221-230、721-730、1331-1340、1521-1530 和 1661-1670 片段对 BJ 正常人包皮成纤维细胞的细胞毒性。结果发现,所有胶原片段都没有细胞毒性。目前正在进一步研究胶原蛋白 IV 片段在再生医学中的潜在用途。
Selection of collagen IV fragments forming the outer sphere of the native protein: Assessment of biological activity for regenerative medicine
The aim of this research was to select the fragments that make up the outer layer of the collagen IV (COL4A6) protein and to assess their potential usefulness for regenerative medicine. It was expected that because protein–protein interactions take place via contact between external domains, the set of peptides forming the outer sphere of collagen IV will determine its interaction with other proteins. Cellulose-immobilized protein fragment libraries treated with polyclonal anti-collagen IV antibodies were used to select the peptides forming the outer sphere of collagen IV. In the first test, 33 peptides that strongly interacted with the polyclonal anti-collagen IV antibodies were selected from a library of non-overlapping fragments of collagen IV. The selected fragments of collagen IV (cleaved from the cellulose matrix) were tested for their cytotoxicity, their effects on cell viability and proliferation, and their impact on the formation of reactive oxygen species (ROS). The studies used RAW 264.7 mouse macrophage cells and Hs 680.Tr human fibroblasts. PrestoBlue, ToxiLight™, and ToxiLight 100% Lysis Control assays were conducted. The viability of fibroblasts cultured with the addition of increasing concentrations of the peptide mix did not show statistically significant differences from the control. Fragments 161–170, 221–230, 721–730, 1331–1340, 1521–1530, and 1661–1670 of COL4A6 were examined for cytotoxicity against BJ normal human foreskin fibroblasts. None of the collagen fragments were found to be cytotoxic. Further research is underway on the potential uses of collagen IV fragments in regenerative medicine.
期刊介绍:
The official Journal of the European Peptide Society EPS
The Journal of Peptide Science is a cooperative venture of John Wiley & Sons, Ltd and the European Peptide Society, undertaken for the advancement of international peptide science by the publication of original research results and reviews. The Journal of Peptide Science publishes three types of articles: Research Articles, Rapid Communications and Reviews.
The scope of the Journal embraces the whole range of peptide chemistry and biology: the isolation, characterisation, synthesis properties (chemical, physical, conformational, pharmacological, endocrine and immunological) and applications of natural peptides; studies of their analogues, including peptidomimetics; peptide antibiotics and other peptide-derived complex natural products; peptide and peptide-related drug design and development; peptide materials and nanomaterials science; combinatorial peptide research; the chemical synthesis of proteins; and methodological advances in all these areas. The spectrum of interests is well illustrated by the published proceedings of the regular international Symposia of the European, American, Japanese, Australian, Chinese and Indian Peptide Societies.