Molecular cloning and functional characterization of bottlenose dolphin (Tursiops truncatus) tumor necrosis factor alpha.
Bottlenose dolphin tumor necrosis factor alpha (doTNF-alpha) cDNA was cloned by reverse transcription polymerase chain reaction (RT-PCR) and the nucleic and deduced amino acid sequences were determined. The sequence of the cDNA clones shows that doTNF-alpha has an open reading frame of 699bp encoding 233 amino acids. The nucleic acid sequence of doTNF-alpha indicates 90, 88, 87, and 79% similarity with the cattle, pig, human, and mouse TNF-alpha gene, respectively. Based on the analysis of human and mouse TNF-alpha molecules, doTNF-alpha is processed to a mature protein with 157 amino acids. The 233 amino acids precursor has a hydrophobic region that could serve as a transmembrane domain. The recombinant doTNF-alpha expressed in Escherichia coli as a glutathione S-transferase fusion protein reacted with anti-human TNF-alpha antibody and exerted cytotoxity to the TNF-alpha sensitive murine cell line L929.
Shoji
Y
Y
Department of Preventive Veterinary Medicine and Animal Health, Nihon University School of Veterinary Medicine, 1866 Kameino, Fujisawa, Kanagawa 252-8510, Japan.
Inoue
Y
Y
Sugisawa
H
H
Itou
T
T
Endo
T
T
Sakai
T
T
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001681:Biological Assay / Q000662:veterinary; D002478:Cells, Cultured; D004247:DNA / Q000737:chemistry; D004289:Dolphins / Q000235:genetics* / Q000276:immunology*; D004926:Escherichia coli / Q000235:genetics; D051379:Mice; D008969:Molecular Sequence Data; D012313:RNA / Q000737:chemistry / Q000302:isolation & purification; D019105:Random Amplified Polymorphic DNA Technique / Q000662:veterinary; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics / Q000633:toxicity; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000662:veterinary; D017386:Sequence Homology, Amino Acid; D014409:Tumor Necrosis Factor-alpha / Q000235:genetics* / Q000502:physiology / Q000633:toxicity
D011994:Recombinant Proteins; D014409:Tumor Necrosis Factor-alpha; D012313:RNA; D004247:DNA