A novel fluorescent protein purified from eel muscle
- 641 Downloads
We discovered that some isolated eel skeletal muscle cells exhibited green fluorescence under a fluorescence stereomicroscope, and we successfully isolated a novel fluorescent protein from the eel muscle homogenate. The protein was a monomer with a molecular mass of 16.5–17 kDa and showed minor and major peaks at 280 and 493 nm, respectively, in the absorption spectrum. The molar extinction coefficient at 493 nm was 41,300 M−1 cm−1 and A280/A493 was 0.083. Excitation and emission spectra of the protein showed maxima at 493 and 527 nm, respectively. Heat treatment at 95°C for 10 min or 5% trichloroacetic acid treatment of the protein caused aggregation of the protein but did not release any fluorescent components such as FAD into the supernatant after centrifugation. Fluorescence of the protein remained after native PAGE, but not after SDS-PAGE. These results indicate that the purified fluorescent protein is not a flavoprotein, and that its fluorescent chromophore is a covalently bound one, such as green fluorescent protein (GFP) from jellyfish Aequoria victoria, but that its fluorescence requires its native conformation within the protein. Based on these results, we can conclude that the fluorescent protein obtained from eel skeletal muscle is a novel GFP-like protein.
KeywordsEel Fluorescent protein GFP GFP-like protein Isolated muscle cells Skeletal muscle
- 2.Hayashi S, Komatsu M (1999) Primary culture of eel hepatocytes—synthesis and secretion of lipoprotein. In: Doyle A, Griffiths JB, Newell DG (eds) Cell and tissue culture: laboratory procedures. Wiley, New York, pp 23A10–23A21Google Scholar
- 3.Kagawa A (1983) Shokuhin seibun-hyo. (Standard tables of food composition in Japan.) In: The Resource Investigation Committee, The Science and Technology Agency, Japan (ed) Shokuhin seibun-hyo. (Standard tables of food composition in Japan, 4th edn.) Kagawa Nutrition University Publishing Division, Tokyo, p 89 (in Japanese)Google Scholar
- 5.Simonetta MP, Pollegioni L, Casalin P, Curti B, Ronchi S (1989) Properties of d-amino-acid oxidase from Rhodotorula gracilis. Eur J Biochem 180:199–204Google Scholar
- 7.Mihalik SJ, McGuinness M, Watkins PA (1991) Purification and characterization of peroxisomal l-pipecolic acid oxidase from monkey liver. J Biol Chem 266:4822–4830Google Scholar