GFP-like chromoproteins as a source of far-red fluorescent proteins

We have employed a new approach to generate novel fluorescent proteins (FPs ) from red absorbing chromoproteins. An identical single amino acid substit ution converted novel chromoproteins from the species Anthozoa (Heteractis crispa, Condylactis gigantea, and Goniopora tenuidens) into far-red FPs (em ission lambda (max) = 615-640 mn). Moreover, coupled site-directed and rand om mutagenesis of the chromoprotein from H. crispa resulted in a unique far -red FP (HcRed) that exhibited bright emission at 645 nm. A clear red shift in fluorescence of HcRed, compared to drFP583 (by more than 60 nm), makes it an ideal additional color for multi-color labeling. Importantly, HcRed i s excitable by 600 nm dye laser, thus promoting new detection channels for multi-color flow cytometry applications. In addition, we generated a dimeri c mutant with similar maturation and spectral properties to tetrameric HcRe d. (C) 2001 Published by Elsevier Science B.V. on behalf of the Federation of European Biochemical Societies.