Very bright green fluorescent proteins from the Pontellid copepod Pontella mimocerami.
BACKGROUND
Fluorescent proteins (FP) homologous to the green fluorescent protein (GFP) from the jellyfish Aequorea victoria have revolutionized biomedical research due to their usefulness as genetically encoded fluorescent labels. Fluorescent proteins from copepods are particularly promising due to their high brightness and rapid fluorescence development.


RESULTS
Here we report two novel FPs from Pontella mimocerami (Copepoda, Calanoida, Pontellidae), which were identified via fluorescence screening of a bacterial cDNA expression library prepared from the whole-body total RNA of the animal. The proteins are very similar in sequence and spectroscopic properties. They possess high molar extinction coefficients (79,000 M(-1) cm(-)) and quantum yields (0.92), which make them more than two-fold brighter than the most common FP marker, EGFP. Both proteins form oligomers, which we were able to counteract to some extent by mutagenesis of the N-terminal region; however, this particular modification resulted in substantial drop in brightness.


CONCLUSIONS
The spectroscopic characteristics of the two P. mimocerami proteins place them among the brightest green FPs ever described. These proteins may therefore become valuable additions to the in vivo imaging toolkit.
Hunt
Marguerite E
ME
Institute for Cellular and Molecular Biology, The University of Texas at Austin, Austin, Texas, United States of America.
Scherrer
Michael P
MP
Ferrari
Frank D
FD
Matz
Mikhail V
MV
D000818:Animals; D033342:Copepoda / Q000378:metabolism*; D005453:Fluorescence; D015723:Gene Library; D049452:Green Fluorescent Proteins / Q000737:chemistry / Q000145:classification* / Q000235:genetics / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D010802:Phylogeny; D055550:Protein Stability
D049452:Green Fluorescent Proteins