International Journal of Molecular Sciences (Mar 2024)

A Combination of Library Screening and Rational Mutagenesis Expands the Available Color Palette of the Smallest Fluorogen-Activating Protein Tag nanoFAST

  • Nadezhda S. Baleeva,
  • Yulia A. Bogdanova,
  • Marina V. Goncharuk,
  • Anatolii I. Sokolov,
  • Ivan N. Myasnyanko,
  • Vadim S. Kublitski,
  • Alexander Yu. Smirnov,
  • Aidar R. Gilvanov,
  • Sergey A. Goncharuk,
  • Konstantin S. Mineev,
  • Mikhail S. Baranov

DOI
https://doi.org/10.3390/ijms25053054
Journal volume & issue
Vol. 25, no. 5
p. 3054

Abstract

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NanoFAST is the smallest fluorogen-activating protein, consisting of only 98 amino acids, used as a genetically encoded fluorescent tag. Previously, only a single fluorogen with an orange color was revealed for this protein. In the present paper, using rational mutagenesis and in vitro screening of fluorogens libraries, we expanded the color palette of this tag. We discovered that E46Q is one of the key substitutions enabling the range of possible fluorogens to be expanded. The introduction of this and several other substitutions has made it possible to use not only orange but also red and green fluorogens with the modified protein.

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