|Characterization of novel orange fluorescent protein cloned from cnidarian tube anemone Cerianthus sp|
|Ip, D.T.-M.; Wong, K.-B.; Wan, D.C.-C. (2007). Characterization of novel orange fluorescent protein cloned from cnidarian tube anemone Cerianthus sp. Mar. Biotechnol. 9(4): 469-478. dx.doi.org/10.1007/s10126-007-9005-5|
|In: Marine Biotechnology. Springer-Verlag: New York. ISSN 1436-2228, more|
Green fluorescent protein; Marine
A novel orange fluorescent protein (OFP) was cloned from the tentacles of Cnidarian tube anemone Cerianthus sp. It consists of 222 amino acid residues with a calculated molecular mass of 25.1 kDa. A BLAST protein sequence homology search revealed that native OFP has 81% sequence identity to Cerianthus membranaceus green fluorescent protein (cmFP512), 38% identity to Entacmaea quadricolor red fluorescent protein (eqFP611), 37% identity to Discosoma red fluorescent protein (DsRed), 36% identity to Fungia concinna Kusabira-orange fluorescent protein (KO), and a mere 21% identity to green fluorescent protein (GFP). It is most likely that OFP also adopts the 11-strand β-barrel structure of fluorescent proteins. Spectroscopic analysis indicated that it has a wide absorption spectrum peak at 548 nm with two shoulders at 487 and 513 nm. A bright orange fluorescence maximum at 573 nm was observed when OFP was excited at 515 nm or above. When OFP was excited well below 515 nm, a considerable amount of green emission maximum at 513 nm was also observed. It has a fluorescence quantum yield (Φ) of 0.64 at 25°C. The molar absorption coefficients (ε) of folded OFP at 278 and 548 nm are 47,000 and 60,000 M-1 ·cm-1, respectively. Its fluorescent brightness (ε Φ) at 25°C is 38,400 M-1 cm-1. Like other orange-red fluorescent proteins, OFP is also tetrameric. It was readily expressed as soluble protein in Escherichia coli at 37°C, and no aggregate was observed in transfected HeLa cells under our experimental conditions. Fluorescent intensity of OFP is detectable over a pH range of 3 to 12.