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Factors Affecting Enzymatic Degradation of Microgel-Bound Peptides
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Pharmacy.
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Pharmacy.
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Pharmacy.
2013 (English)In: Biomacromolecules, ISSN 1525-7797, E-ISSN 1526-4602, Vol. 14, no 7, 2317-2325 p.Article in journal (Refereed) Published
Abstract [en]

Proteolytic degradation and release of microgel-bound peptides was investigated for trypsin, poly(acrylic acid-co-acrylamide) microgels (70-90 mu m in diameter), and oppositely charged polylysine, using a method combination of confocal microscopy and micromanipulator-assisted light microscopy. Results show that trypsin-induced release of polylysine increased with increasing trypsin concentration, decreasing microgel charge density and decreasing peptide molecular weight. While the microgel offered good protection against enzymatic degradation at high microgel charge density, it was also observed that the cationic peptide enabled trypsin to bind throughout the peptide-loaded microgels, even when it did not bind to the peptide-void ones. With the exception of highly charged microgels, proteolytic degradation throughout the peptide-loaded microgel resulted in the generation of short and non-adsorbing peptide stretches, giving rise to the concentration and peptide length dependence observed. A simple random scission model was able to qualitatively capture these experimental findings. collectively, the results demonstrate that microgel charge density, peptide molecular weight, and enzyme concentration greatly influence degradation/release of microgel-bound peptides and need to be considered in the use of microgels, e.g., as carriers for protein and peptide drugs.

Place, publisher, year, edition, pages
2013. Vol. 14, no 7, 2317-2325 p.
National Category
Natural Sciences
Identifiers
URN: urn:nbn:se:uu:diva-204977DOI: 10.1021/bm400431fISI: 000321793700021OAI: oai:DiVA.org:uu-204977DiVA: diva2:641369
Available from: 2013-08-16 Created: 2013-08-13 Last updated: 2017-12-06Bibliographically approved
In thesis
1. Microgel Interactions with Peptides and Proteins: Consequence of Peptide and Microgel Properties
Open this publication in new window or tab >>Microgel Interactions with Peptides and Proteins: Consequence of Peptide and Microgel Properties
2015 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Microgels are lightly cross-linked hydrogel particles in the sub-micrometer to micrometer size range with a capacity to drastically change their volume in response to changes in the external environment. Microgels have an ability to bind and store substances such as biomacromolecular drugs, notably proteins and peptides, and release them upon stimuli, making them potential candidates as drug delivery vehicles and functional biomaterials. This thesis aims at clarifying important factors affecting peptide-microgel interactions. These interactions were studied by micromanipulator-assisted light and fluorescence microscopy focusing on microgel deswelling in response to peptide binding, as well as re-swelling in response to peptide release or enzymatic degradation. To evaluate peptide uptake in microgels, solution depletion measurements were used whereas the peptide secondary structure was investigated by circular dichroism. In addition, the peptide and enzyme distribution within microgels was analyzed with confocal microscopy.

Results presented in this thesis demonstrate that peptide incorporation into microgels, as well as peptide-induced microgel deswelling, increases with peptide length and charge density. In addition, results demonstrate that the peptide charge (length) rather than peptide charge density determines microgels deswelling. End-to-end cyclization is shown to not noticeably influence peptide-microgel interactions, suggesting that peptide cyclization can be used in combination with oppositely charged microgel carriers to improve the proteolytic and chemical stability of the peptide compared to the corresponding linear variant. Peptide secondary structure is found to drastically affect peptide incorporation into, and release from, oppositely charged microgels. Furthermore, it is shown that microgel charge density, peptide molecular weight, and enzyme concentration all greatly influence microgel bound peptide degradation. Of importance for applications, protective effects of microgels against proteolytic peptide degradation are observed only at sufficiently high microgel charge densities. Enzyme-mediated microgel degradation is shown to increase with increasing enzyme concentration, while an increased peptide loading in microgels causes a concentration-dependent decrease in microgel degradation.

Taken together, results obtained in this work have provided some insight into factors of importance for rational use of microgels as delivery systems for protein or peptide drugs, but also in a host of other biomedical applications using weakly cross-linked polymer systems.

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2015. 65 p.
Series
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Pharmacy, ISSN 1651-6192 ; 196
Keyword
Binding, Degradation, Enzyme, Gel, Hydrogel, Microgel, Peptide, Protein, Release
National Category
Pharmaceutical Sciences Physical Chemistry Materials Engineering
Research subject
Pharmaceutical Physical Chemistry
Identifiers
urn:nbn:se:uu:diva-242893 (URN)978-91-554-9157-4 (ISBN)
Public defence
2015-03-20, B21, BMC, Husargatan 3, Uppsala, 09:15 (English)
Opponent
Supervisors
Available from: 2015-02-24 Created: 2015-02-02 Last updated: 2015-03-11

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Mansson, RonjaFrenning, GöranMalmsten, Martin

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