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Circular Permutation of Red Fluorescent Proteins

By Bo Shui, Qi Wang, Frank Lee, Laura J. Byrnes, Dmitry M. Chudakov, Sergey A. Lukyanov, Holger Sondermann and Michael I. Kotlikoff

Abstract

Circular permutation of fluorescent proteins provides a substrate for the design of molecular sensors. Here we describe a systematic exploration of permutation sites for mCherry and mKate using a tandem fusion template approach. Circular permutants retaining more than 60% (mCherry) and 90% (mKate) brightness of the parent molecules are reported, as well as a quantitative evaluation of the fluorescence from neighboring mutations. Truncations of circular permutants indicated essential N- and C- terminal segments and substantial flexibility in the use of these molecules. Structural evaluation of two cp-mKate variants indicated no major conformational changes from the previously reported wild-type structure, and cis conformation of the chromophores. Four cp-mKates were identified with over 80% of native fluorescence, providing important new building blocks for sensor and complementation experiments

Topics: Research Article
Publisher: Public Library of Science
OAI identifier: oai:pubmedcentral.nih.gov:3103546
Provided by: PubMed Central

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