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Magnetic catechol-chitosan with bioinspired adhesive surface: preparation and immobilization of ?-transaminase.

ABSTRACT: The magnetic chitosan nanocomposites have been studied intensively and been used practically in various biomedical and biological applications including enzyme immobilization. However, the loading capacity and the remained activity of immobilized enzyme based on existing approaches are not satisfied. Simpler and more effective immobilization strategies are needed. Here we report a simple catechol modified protocol for preparing a novel catechol-chitosan (CCS)-iron oxide nanoparticles (IONPs) composites carrying adhesive moieties with strong surface affinity. The ?-transaminase (?-TA) was immobilized onto this magnetic composite via nucleophilic reactions between catechol and ?-TA. Under optimal conditions, 87.5% of the available ?-TA was immobilized on the composite, yielding an enzyme loading capacity as high as 681.7 mg/g. Furthermore, the valuation of enzyme activity showed that ?-TA immobilized on CCS-IONPs displayed enhanced pH and thermal stability compared to free enzyme. Importantly, the immobilized ?-TA retained more than 50% of its initial activity after 15 repeated reaction cycles using magnetic separation and 61.5% of its initial activity after storage at 4°C in phosphate buffered saline (PBS) for 15 days. The results suggested that such adhesive magnetic composites may provide an improved platform technology for bio-macromolecules immobilized.

SUBMITTER: Ni K

PROVIDER: S-EPMC3398885 | biostudies-literature | 2012

REPOSITORIES: biostudies-literature

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Magnetic catechol-chitosan with bioinspired adhesive surface: preparation and immobilization of ω-transaminase.

Ni Kefeng K Zhou Xu X Zhao Li L Wang Hualei H Ren Yuhong Y Wei Dongzhi D

PloS one 20120717 7

The magnetic chitosan nanocomposites have been studied intensively and been used practically in various biomedical and biological applications including enzyme immobilization. However, the loading capacity and the remained activity of immobilized enzyme based on existing approaches are not satisfied. Simpler and more effective immobilization strategies are needed. Here we report a simple catechol modified protocol for preparing a novel catechol-chitosan (CCS)-iron oxide nanoparticles (IONPs) com ...[more]

PMID: 22815930

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Project description:High-fidelity surface functional group (e.g., N-hydroxysuccinimide (NHS) reactive ester) patterning is readily and reliably achieved on commercial poly(ethylene glycol) (PEG)-based polymer films already known to exhibit high performance non-fouling properties in full serum and in cell culture conditions. NHS coupling chemistry co-patterned with methoxy-capped PEG using photolithographic methods is directly spatially imaged using imaging time-of-flight secondary ion mass spectrometry (ToF-SIMS) and principal components statistical analysis. Patterned NHS surface reactive zones are clearly resolved at high sensitivity despite the complexity of the polymer matrix chemistry. ToF-SIMS imaging also reveals the presence of photo-resist residue remaining from typical photolithography processing methods. High cross-correlation between various ion-derived ToF-SIMS images is observed, providing sensitive chemical corroboration of pattern chemistry and biological reactivity in complex milieu. Surface-specific protein coupling is observed first by site-selective reaction of streptavidin with NHS patterns, followed by identical patterns of biotinylated Alexa-labeled albumin coupling. This suggests that streptavidin immobilized on the patterns remains bioactive. Fluorescently labeled full serum is shown to react selectively with NHS-reactive regions, with minimal signal from methoxy-capped regions. Insufficient serum is adsorbed under any conditions to these surfaces to support cell attachment in serum-containing media. This reflects the high intrinsic non-adsorptive nature of this chemistry. Fibroblasts attach and proliferate in serum culture only when a cell adhesion peptide (RGD) is first grafted to NHS regions on the PEG-based surfaces. Longer-term serum-based cell culture retains high cell-pattern fidelity that correlates with chemical imaging of both the NHS and RGD patterns and also lack of cell adhesion to methoxy-capped regions. Cell staining shows orientation of adherent cells within the narrow patterned areas. Cell patterns are consistently retained beyond 15 days in serum media.

Dataset Information

Magnetic catechol-chitosan with bioinspired adhesive surface: preparation and immobilization of ?-transaminase.

Publications

Magnetic catechol-chitosan with bioinspired adhesive surface: preparation and immobilization of ω-transaminase.

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