very bright green fluorescent proteins from the pontellid copepod pontella mimocerami非常明亮的绿色荧光蛋白的pontellid桡足动物pontella mimocerami.pdfVIP

Very Bright Green Fluorescent Proteins from the Pontellid Copepod Pontella mimocerami 1 1 2 3 Marguerite E. Hunt , Michael P. Scherrer , Frank D. Ferrari , Mikhail V. Matz * 1 Institute for Cellular and Molecular Biology, The University of Texas at Austin, Austin, Texas, United States of America, 2 Department of Invertebrate Zoology, National Museum of Natural History, Smithsonian Institution, Suitland, Maryland, United States of America, 3 Section of Integrative Biology, The University of Texas at Austin, Austin, Texas, United States of America Abstract 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 M21 cm2) 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. mi