RESUMEN
BACKGROUND: Superparamagnetic iron oxide nanoparticles (MNP) offer several advantages for applications in biomedical and biotechnological research. In particular, MNP-based immobilization of enzymes allows high surface-to-volume ratio, good dispersibility, easy separation of enzymes from the reaction mixture, and reuse by applying an external magnetic field. In a biotechnological perspective, extremophilic enzymes hold great promise as they often can be used under non-conventional harsh conditions, which may result in substrate transformations that are not achievable with normal enzymes. This prompted us to investigate the effect of MNP bioconjugation on the catalytic properties of a thermostable carboxypeptidase from the hyperthermophilic archaeon Sulfolobus solfataricus (CPSso), which exhibits catalytic properties that are useful in synthetic processes. RESULTS: CPSso was immobilized onto silica-coated iron oxide nanoparticles via NiNTA-His tag site-directed conjugation. Following the immobilization, CPSso acquired distinctly higher long-term stability at room temperature compared to the free native enzyme, which, in contrast, underwent extensive inactivation after 72 h incubation, thus suggesting a potential utilization of this enzyme under low energy consumption. Moreover, CPSso conjugation also resulted in a significantly higher stability in organic solvents at 40°C, which made it possible to synthesize N-blocked amino acids in remarkably higher yields compared to those of free enzyme. CONCLUSIONS: The nanobioconjugate of CPSso immobilized on silica-coated magnetic nanoparticles exhibited enhanced stability in aqueous media at room temperature as well as in different organic solvents. The improved stability in ethanol paves the way to possible applications of immobilized CPSso, in particular as a biocatalyst for the synthesis of N-blocked amino acids. Another potential application might be amino acid racemate resolution, a critical and expensive step in chemical synthesis.
Asunto(s)
Carboxipeptidasas/química , Enzimas Inmovilizadas/química , Nanoconjugados/química , Sulfolobus solfataricus/enzimología , Estabilidad de Enzimas , Compuestos Férricos/química , Dióxido de Silicio/químicaRESUMEN
AIM: We investigate MAdCAM-1 as a reliable target to detect active bowel inflammation for selective noninvasive nanodiagnostics. MATERIALS & METHODS: We coupled anti-MAdCAM-1 antibodies to manganese oxide nanoparticles, and analyzed nanoconjugate biodistribution and safety in murine model of inflammatory bowel disease by imaging and histology. RESULTS: Nanoparticles were stable and nontoxic. Upon administration in colitic mice, anti-MAdCAM-1 functionalized nanoparticles preferentially localized in the inflamed bowel, whereas untargeted nanoparticles were more rapidly washed out. Nanoparticles did not induce lesions in nontarget organs. CONCLUSION: Anti-MAdCAM-1 functionalized nanoparticles detected active bowel inflammation foci, accurately following MAdCAM-1 expression pattern. These nanoconjugates could be a promising noninvasive imaging system for an early and accurate follow-up in patients affected by acute colitis.
Asunto(s)
Anticuerpos/química , Colitis/diagnóstico por imagen , Inmunoglobulinas/inmunología , Compuestos de Manganeso/química , Nanopartículas del Metal/química , Mucoproteínas/inmunología , Óxidos/química , Animales , Moléculas de Adhesión Celular , Supervivencia Celular , Colitis/metabolismo , Colitis/patología , Hemólisis , Humanos , Inmunoglobulinas/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Mucoproteínas/metabolismo , Tamaño de la Partícula , Propiedades de Superficie , Distribución TisularRESUMEN
AIMS: Chronic lung allograft dysfunction represents the main cause of death after lung transplantation, and so far there is no effective therapy. Mesenchymal cells (MCs) are primarily responsible for fibrous obliteration of small airways typical of chronic lung allograft dysfunction. Here, we engineered gold nanoparticles containing a drug in the hydrophobic section to inhibit MCs, and exposing on the outer hydrophilic surface a monoclonal antibody targeting a MC-specific marker (half-chain gold nanoparticles with everolimus). MATERIALS & METHODS: Half-chain gold nanoparticles with everolimus have been synthesized and incubated with MCs to evaluate the effect on proliferation and apoptosis. RESULTS & DISCUSSION: Drug-loaded gold nanoparticles coated with the specific antibody were able to inhibit proliferation and induce apoptosis without stimulating an inflammatory response, as assessed by in vitro experiments. CONCLUSION: These findings demonstrate the effectiveness of our nanoparticles in inhibiting MCs and open new perspectives for a local treatment of chronic lung allograft dysfunction.