Journal
COLLOIDS AND SURFACES B-BIOINTERFACES
Volume 171, Issue -, Pages 351-357Publisher
ELSEVIER
DOI: 10.1016/j.colsurfb.2018.07.047
Keywords
Polymer brushes; Enzymes; Biocatalysis; Click chemistry
Funding
- Leibniz Institute of Polymer Research Dresden
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Thermoresponsive poly(N-isopropylacrylamide) (PNIPAAm) brushes with terminal click functionality can be used to selectively immobilize enzymes. Exploiting their inherent protein-repellent and thus non-fouling properties, surfaces with specific bioactivity can be created in this way. This report describes the functionalization of alkyne-PNIPAAm brushes with two biotin linkers of different poly(ethylene glycol) (PEG) spacer length via click chemistry and the subsequent immobilization of streptavidin-conjugated horseradish peroxidase (SA-HRP) by the strong interaction between biotin and streptavidin. Spectroscopic ellipsometry is used to quantify the biotin and the SA-HRP amount on the PNIPAAm brushes. Enzyme activities are determined by UV-vis spectroscopy. A better accessibility to the alkyne-functionalized chain ends for the short biotin-PEG(3) linker leads to a higher biotin amount on these PNIPAAm brushes, which in turn results in higher SA-HRP amounts on biotin-PEG(3)-modified brushes in the swollen (20 degrees C) and collapsed state (37 degrees C) compared to biotin-PEG(23)-modified ones. For both linkers, an increased immobilization temperature leads to higher SA-HRP amounts due to an enhanced reaction kinetics and mobility. All immobilized SA-HRP amounts are in range of 81-98% monolayer coverage, except for SA-HRP immobilized on biotin-PEG(23)-modified PNIPAAm brushes at 20 degrees C, exhibiting only 43% monolayer coverage. A high mobility of the biotin linker combined with a low surface loading of SA-HRP is found to be beneficial for the activity of SA-HRP. Hence, the highest specific activity as measured by HRP-catalyzed 3,3',5,5'-tetramethylbenzidine (TMB) oxidation is found for a low SA-HRP amount on biotin-PEG(23)-modified brushes.
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