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Utilizing split-NanoLuc luciferase fragments as luminescent probes for protein solubility in living cells.

ABSTRACT: Protein misfolding and aggregation is now recognized as a hallmark of numerous human diseases. Standard bioanalytical techniques for monitoring protein aggregation generally rely on small molecules that provide an optical readout of fibril formation. While these methods have been useful for mechanistic studies, additional approaches are required to probe the equilibrium between soluble and insoluble protein within living systems. Such approaches could provide platforms for the identification of inhibitors of protein aggregation as well as a means to investigate the effect of mutations on protein aggregation in model systems. In this chapter, we provide detailed protocols for employing split-NanoLuc luciferase (Nluc) fragments to monitor changes in protein solubility in bacterial and mammalian cells. This sensitive luminesce-based assay can report upon changes in protein solubility induced by inhibitors and disease-relevant mutations.

SUBMITTER: Nelson TJ

PROVIDER: S-EPMC6567984 | biostudies-literature | 2019

REPOSITORIES: biostudies-literature

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Utilizing split-NanoLuc luciferase fragments as luminescent probes for protein solubility in living cells.

Nelson Travis J TJ Zhao Jia J Stains Cliff I CI

Methods in enzymology 20190309

Protein misfolding and aggregation is now recognized as a hallmark of numerous human diseases. Standard bioanalytical techniques for monitoring protein aggregation generally rely on small molecules that provide an optical readout of fibril formation. While these methods have been useful for mechanistic studies, additional approaches are required to probe the equilibrium between soluble and insoluble protein within living systems. Such approaches could provide platforms for the identification of ...[more]

PMID: 31155065

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