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Quantitative real-time kinetics of optogenetic proteins CRY2 and CIB1/N using single-molecule tools.

ABSTRACT: In this work we evaluate the interaction of two optogenetic protein variants (CIB1, CIBN) with their complementary protein CRY2 by single-molecule tools in cell-free extracts. After validating the blue light induced co-localization of CRY2 and CIB1/N by Förster resonance energy transfer (FRET) in live cells, a fluorescence correlation spectroscopy (FCS) based method was developed to quantitatively determine the in vitro association of the extracted proteins. Our experiments suggest that CIB1, in comparison with CIBN, possesses a better coupling efficiency with CRY2 due to its intact protein structure and lower diffusion rate within 300s detection window.

SUBMITTER: Cui Y

PROVIDER: S-EPMC4341968 | biostudies-literature | 2014 Aug

REPOSITORIES: biostudies-literature

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Quantitative real-time kinetics of optogenetic proteins CRY2 and CIB1/N using single-molecule tools.

Cui Yi Y Choudhury Samrat Roy SR Irudayaraj Joseph J

Analytical biochemistry 20140426

In this work we evaluate the interaction of two optogenetic protein variants (CIB1, CIBN) with their complementary protein CRY2 by single-molecule tools in cell-free extracts. After validating the blue light induced co-localization of CRY2 and CIB1/N by Förster resonance energy transfer (FRET) in live cells, a fluorescence correlation spectroscopy (FCS) based method was developed to quantitatively determine the in vitro association of the extracted proteins. Our experiments suggest that CIB1, in ...[more]

PMID: 24780222

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