Engineering and Characterization of 3-Aminotyrosine-Derived Red Fluorescent Variants of Circularly Permutated Green Fluorescent Protein.

ABSTRACT: Introducing 3-aminotyrosine (aY), a noncanonical amino acid (ncAA), into green fluorescent protein (GFP)-like chromophores shows promise for achieving red-shifted fluorescence. However, inconsistent results, including undesired green fluorescent species, hinder the effectiveness of this approach. In this study, we optimized expression conditions for an aY-derived cpGFP (aY-cpGFP). Key factors like rich culture media and oxygen restriction pre- and post-induction enabled high-yield, high-purity production of the red-shifted protein. We also engineered two variants of aY-cpGFP with enhanced brightness by mutating a few amino acid residues surrounding the chromophore. We further investigated the sensitivity of the aY-derived protein to metal ions, reactive oxygen species (ROS), and reactive n

SUBMITTER: Zhang H 

PROVIDER: S-EPMC10813706 | biostudies-literature | 2024 Jan

REPOSITORIES: biostudies-literature