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
Publisher: Public Library of Science
Date Published: 27-May-2011
Author(s): Shui B., Wang Q., Lee F., Byrnes L., Chudakov D., Lukyanov S., Sondermann H., Kotlikoff M.