Chemical and Biomolecular Engineering - Theses
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ItemSpider silk-inspired functional materials with tailored surface properties for biomedical applications( 2022)Biomaterials science is an increasingly important and constantly evolving field of science. Only intensive cooperation between different disciplines and a deep understanding of the physical and chemical interactions within developed materials and the biological system as a whole lead to the successful development of new biomaterials. Biocompatibility plays a central role here. It must be possible to assess whether the material is compatible with the respective application, e.g., implantation in hard or soft tissue. Here, a further distinction can be made between structural and surface compatibility. Structural compatibility covers the structure, shape, and mechanical property interactions in a biological environment. Surface compatibility summarizes the adaptation of chemical, physical, biological, and morphological surface properties to the biological environment. Consequently, the surface properties of a biomaterial are crucial for its biocompatibility and interactions with the host system. Materials used as biomaterials must fulfill a wide range of requirements. They should have excellent mechanical stability, be biocompatible and, depending on the requirements, bioinert or bioactive. For example, bioactive biomaterials are used to increase or control interaction with cells. Synthetic polymers usually have excellent mechanical properties but then lack biocompatibility, whereas natural polymers often have excellent biocompatibility but then are mechanically very weak and therefore not suitable for applications with high mechanical stress. A promising material that exhibits the advantages of both classes of polymers is spider silk. Spider silk has been used since ancient times as wound dressings and suture material as it is mechanically resilient and elastic and elicits little to no immune response. Natural spider silk cannot be used as a biomaterial on a large scale due to the cannibalistic behavior of most spider species and changing quality of silk. Therefore, this work presents two approaches utilizing materials inspired from natural spider silk to create functional, modifiable, mechanically resilient, and biocompatible coatings. The first approach is bioengineered recombinant spider silk proteins. Before biotechnological production, these proteins produced can be genetically modified in E.Coli bacteria. In this work, twelve different spider silk protein variants are used and investigated concerning their biocompatibility, biodegradability, and interaction with proteins, cells, and human blood. These spider silk protein variants are non-toxic and can be resorbed by the body as they consist solely of amino acids. The second approach is based on synthetic polypeptides prepared by the continuous assembly polymerization (CAP) method, published for the first time, using reversible-addition-fragmentation chain-transfer (RAFT) polymerization, or CAP-RAFT. Polypeptides were selected based on amino acids found in natural spider silk (L-lysine and L-glutamic acid). These coatings based on synthetic polypeptides were investigated concerning secondary structure and biodegradability. CAP-RAFT was established as a viable strategy to prepare surface-limited cross-linked polypeptide films with precise film thickness control and novel properties such as specific secondary structure formation and biodegradation. This variability of secondary structure combined with enzymatic degradation shows high potential for numerous biological applications. In the present work, secondary structure formation and assembly of the spider silk-inspired materials on coatings were investigated in detail. Firstly, the effect of coating thickness on the structural properties (beta-sheet fraction) was investigated from the nanoscale to the microscale. A coating thickness-dependent assembly and phase separation model is presented. In addition, the orientation of beta-sheets in recombinant spider silk coatings was investigated. Another important aspect of surface biocompatibility is the structure-property relationship of these spider silk-based materials. Concerning applications in the biomedical field, the interaction between material and biological environment is essential. Several aspects are studied in detail: specifically surface charge, surface chemistry, surface topography, and surface hydrophilicity. These aspects were analyzed to understand the interaction with proteins, cells, and blood as well as their biodegradability. Based on the results of the respective studies, it was possible to categorize the different spider silk variants into bioinert and bioactive variants and assign their subsequent potential biomedical applications. Positively charged spider silk protein variants are bioactive and have the most significant interaction with cells and blood. Modification with the cell-binding peptide improved cell adhesion of all variants used. Amino acid sequences based on the natural Araneus diadematus fibroin (ADF) 3 protein showed significantly faster enzymatic degradation than the protein variants based on the amino acid sequence of ADF4. The introduction of three-dimensional patterns on the coating surface can significantly increase the adhesion of cells to material (negatively charged variant), which shows little adhesion of cells as a smooth coating. In this dissertation, the structure formation, assembly, and structure-property relationships of spider silk-inspired materials were systematically investigated. These spider silk-inspired materials possessed a high potential for application in various biomedicine fields due to the diverse modification possibilities in terms of morphology, amino acid sequence, and charge.