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Droplet-Based Microfluidic Temperature-Jump Platform for the Rapid Assessment of Biomolecular Kinetics

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YANG Tianjin VILLOIS Alessia KUNKA Antonín GRIGOLATO Fulvio AROSIO Paolo PROKOP Zbyněk DEMELLO Andrew STAVRAKIS Stavros

Rok publikování 2022
Druh Článek v odborném periodiku
Časopis / Zdroj Analytical chemistry
Fakulta / Pracoviště MU

Přírodovědecká fakulta

Citace
www https://doi.org/10.1021/acs.analchem.2c03009
Doi http://dx.doi.org/10.1021/acs.analchem.2c03009
Klíčová slova Biotechnology; Fluid dynamics; Kinetics; Liquids; Nanofibers
Popis Protein folding, unfolding, and aggregation are important in a variety of biological processes and intimately linked to “protein misfolding diseases”. The ability to perform experiments at different temperatures allows the extraction of important information regarding the kinetics and thermodynamics of such processes. Unfortunately, conventional stopped-flow methods are difficult to implement, generate limited information, and involve complex sample handling. To address this issue, we present a temperature-controlled droplet-based microfluidic platform that allows measurement of reaction kinetics on millisecond to second timescales and at temperatures between ambient and 90 °C. The utility of the microfluidic platform for measuring fast biomolecular kinetics at high temperatures is showcased through the investigation of the unfolding kinetics of haloalkane dehalogenases and the elongation of fibrils composed of the amyloid ß peptide associated with Alzheimer’s disease. In addition, a deep-ultraviolet (UV) fluorescence microscope was developed for the on-chip recording of protein intrinsic fluorescence spectrum originating from aromatic amino acid residues. We envision that the developed optofluidic platform will find wide applicability in the analysis of biological processes, such as protein refolding and phase separation.
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