New antimicrobial biomaterials based on recombinant spider silks
Spider silk is a protein-based material with exceptional mechanical properties together with low immunogenicity and pyrogenicity which makes it useful in biomedical applications. The cannibalistic and highly territorial nature of most spiders prevents high-density farming; therefore, the availabilit...
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| Format: | Thesis (University of Nottingham only) |
| Language: | English |
| Published: |
2022
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| Subjects: | |
| Online Access: | https://eprints.nottingham.ac.uk/68429/ |
| _version_ | 1848800487086751744 |
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| author | Beinarovica, Jolanta |
| author_facet | Beinarovica, Jolanta |
| author_sort | Beinarovica, Jolanta |
| building | Nottingham Research Data Repository |
| collection | Online Access |
| description | Spider silk is a protein-based material with exceptional mechanical properties together with low immunogenicity and pyrogenicity which makes it useful in biomedical applications. The cannibalistic and highly territorial nature of most spiders prevents high-density farming; therefore, the availability of their silk in a usable form is very limited. Recombinant production of silks is explored as an alternative mean of production. This study uses two miniature recombinant silk proteins (mini-spidroins) with distinct biochemical nature -NT2RepCT and 4RepCT - to create new materials for drug delivery.
A procedure for a column-free, scale up compatible purification of highly water soluble spidroin NT2RepCT has been developed. NT2RepCT was then processed into a colloidal drug delivery system that could be loaded with a model drug and exhibited a pH-dependent controlled drug release profile. It was found that NT2RepCT particles have a polydisperse size range on a micron scale, and that they are unstable in water, which makes them useful for in situ and temporary embolic applications.
Further, the 4RepCT3Aha mini-spidroin was expressed and purified. In this construct, each methionine residue is replaced with a synthetic methionine analogue L-azidohomoalanine (Aha) that carries a terminal azide in its side chain. The azide acts as a selectively chemically reactive, bioorthogonal group for bioconjugations using copper-catalysed azide-alkyne cycloaddition (CuAAC) knows as the “click reaction”. Using this methodology, a selection of antimicrobial ligands (triclosan, chloramphenicol, ciprofloxacin, erythromycin, levofloxacin, and nitroxoline) bearing a labile linker with a terminal alkyne were conjugated to 4RepCT3Aha , creating a library of antimicrobial conjugates. The resulting conjugates were processed into films that showed significant antimicrobial activity against the Gram-negative Escherichia coli (E. coli) and the Gram-positive Staphylococcus aureus (S. aureus) in a novel plate-based, high throughput-compatible assay. Further, it was found that ‘clickable’ antimicrobial ligands could also be conjugated to pre-formed Aha-bearing silk films. When the antimicrobial ligands were conjugated to pre-formed films, their biocidal activity was lower than that of films made from soluble 4RepCT3Aha -ligand conjugates but the activity was statistically significant compared to films dipped in antibiotic solution.
In addition, the 4RepCT3Aha mini-spidroin was functionalised with quaternary ammonium-based ligands via a non-labile linker prior to processing of the conjugate into a surface coating. In this approach, a cationic, contact-active antimicrobial surface was created that showed significant antimicrobial effect against E. coli in a range of conventional microbiological assays. For this material, a tailored high-throughput compatible assay to analyse the metabolic activity and biomass increase of surface-adherent bacteria was developed. In this assay, it was found that quaternary-ammonium bearing ligands have activity against E. coli, but not S. aureus or Pseudomonas aeruginosa.
In conclusion, this work describes a range of new antimicrobial materials based on miniature spider silks that can serve as a drug delivery vehicle in different biomedically relevant scenarios. By combining silk’s uniquely biocompatible nature with a tailored functionality and modifiable release kinetics, these novel biomaterials are promising candidates for drug delivery applications. |
| first_indexed | 2025-11-14T20:52:20Z |
| format | Thesis (University of Nottingham only) |
| id | nottingham-68429 |
| institution | University of Nottingham Malaysia Campus |
| institution_category | Local University |
| language | English |
| last_indexed | 2025-11-14T20:52:20Z |
| publishDate | 2022 |
| recordtype | eprints |
| repository_type | Digital Repository |
| spelling | nottingham-684292025-02-28T15:14:45Z https://eprints.nottingham.ac.uk/68429/ New antimicrobial biomaterials based on recombinant spider silks Beinarovica, Jolanta Spider silk is a protein-based material with exceptional mechanical properties together with low immunogenicity and pyrogenicity which makes it useful in biomedical applications. The cannibalistic and highly territorial nature of most spiders prevents high-density farming; therefore, the availability of their silk in a usable form is very limited. Recombinant production of silks is explored as an alternative mean of production. This study uses two miniature recombinant silk proteins (mini-spidroins) with distinct biochemical nature -NT2RepCT and 4RepCT - to create new materials for drug delivery. A procedure for a column-free, scale up compatible purification of highly water soluble spidroin NT2RepCT has been developed. NT2RepCT was then processed into a colloidal drug delivery system that could be loaded with a model drug and exhibited a pH-dependent controlled drug release profile. It was found that NT2RepCT particles have a polydisperse size range on a micron scale, and that they are unstable in water, which makes them useful for in situ and temporary embolic applications. Further, the 4RepCT3Aha mini-spidroin was expressed and purified. In this construct, each methionine residue is replaced with a synthetic methionine analogue L-azidohomoalanine (Aha) that carries a terminal azide in its side chain. The azide acts as a selectively chemically reactive, bioorthogonal group for bioconjugations using copper-catalysed azide-alkyne cycloaddition (CuAAC) knows as the “click reaction”. Using this methodology, a selection of antimicrobial ligands (triclosan, chloramphenicol, ciprofloxacin, erythromycin, levofloxacin, and nitroxoline) bearing a labile linker with a terminal alkyne were conjugated to 4RepCT3Aha , creating a library of antimicrobial conjugates. The resulting conjugates were processed into films that showed significant antimicrobial activity against the Gram-negative Escherichia coli (E. coli) and the Gram-positive Staphylococcus aureus (S. aureus) in a novel plate-based, high throughput-compatible assay. Further, it was found that ‘clickable’ antimicrobial ligands could also be conjugated to pre-formed Aha-bearing silk films. When the antimicrobial ligands were conjugated to pre-formed films, their biocidal activity was lower than that of films made from soluble 4RepCT3Aha -ligand conjugates but the activity was statistically significant compared to films dipped in antibiotic solution. In addition, the 4RepCT3Aha mini-spidroin was functionalised with quaternary ammonium-based ligands via a non-labile linker prior to processing of the conjugate into a surface coating. In this approach, a cationic, contact-active antimicrobial surface was created that showed significant antimicrobial effect against E. coli in a range of conventional microbiological assays. For this material, a tailored high-throughput compatible assay to analyse the metabolic activity and biomass increase of surface-adherent bacteria was developed. In this assay, it was found that quaternary-ammonium bearing ligands have activity against E. coli, but not S. aureus or Pseudomonas aeruginosa. In conclusion, this work describes a range of new antimicrobial materials based on miniature spider silks that can serve as a drug delivery vehicle in different biomedically relevant scenarios. By combining silk’s uniquely biocompatible nature with a tailored functionality and modifiable release kinetics, these novel biomaterials are promising candidates for drug delivery applications. 2022-03-15 Thesis (University of Nottingham only) NonPeerReviewed application/pdf en cc_by https://eprints.nottingham.ac.uk/68429/1/Beinarovica%20Jolanta%2014304137%20PhD%20Chemistry.pdf Beinarovica, Jolanta (2022) New antimicrobial biomaterials based on recombinant spider silks. PhD thesis, University of Nottingham. spider silk antimicrobial biomaterials drug delivery systems |
| spellingShingle | spider silk antimicrobial biomaterials drug delivery systems Beinarovica, Jolanta New antimicrobial biomaterials based on recombinant spider silks |
| title | New antimicrobial biomaterials based on recombinant spider silks |
| title_full | New antimicrobial biomaterials based on recombinant spider silks |
| title_fullStr | New antimicrobial biomaterials based on recombinant spider silks |
| title_full_unstemmed | New antimicrobial biomaterials based on recombinant spider silks |
| title_short | New antimicrobial biomaterials based on recombinant spider silks |
| title_sort | new antimicrobial biomaterials based on recombinant spider silks |
| topic | spider silk antimicrobial biomaterials drug delivery systems |
| url | https://eprints.nottingham.ac.uk/68429/ |