title: Molecular cloning and expression of α-globin and β-globin genes from crocodile (Crocodylus siamensis).
abstract: The first report of complete nucleotide sequences for α- and β-globin chains from the Siamese hemoglobin (Crocodylus siamensis) is given in this study. The cDNAs encoding α- and β-globins were cloned by RT-PCR using the degenerate primers and by the rapid amplification of cDNA ends method. The full-length α-globin cDNA contains an open reading frame of 423 nucleotides encoding 141 amino acid residues, whereas the β-globin cDNA contains an open reading frame of 438 nucleotides encoding 146 amino acid residues. The authenticity of both α- and β-globin cDNA clones were also confirmed by the heterologous expression in Escherichia coli (E. coli). This is the first time that the recombinant C. siamensis globins were produced in prokaryotic system. Additionally, the heme group was inserted into the recombinant proteins and purified heme-bound proteins were performed by affinity chromatography using Co(2+)-charged Talon resins. The heme-bound proteins appeared to have a maximum absorbance at 415 nm, indicated that the recombinant proteins bound to oxygen and formed active oxyhemoglobin (HbO2). The results indicated that recombinant C. siamensis globins were successfully expressed in prokaryotic system and possessed an activity as ligand binding protein.
authors: Protein and Proteomics Research Center for Commercial and Industrial Purposes (ProCCI), Department of Biochemistry, Faculty of Science, Khon Kaen University, Khon Kaen, 40002, Thailand.
authors: Preeyanan
authors: P
authors: Anwised
authors: Thai
authors: T
authors: Kabbua
authors: Theeranan
authors: T
authors: Temsiripong
authors: Apisak
authors: A
authors: Dhiravisit
authors: Sarawut
authors: S
authors: Jitrapakdee
authors: Tomohiro
authors: T
authors: Araki
authors: Kazunari
authors: K
authors: Yoneda
authors: Sompong
authors: S
authors: Thammasirirak
mesh_terms: D000489:Alligators and Crocodiles / Q000235:genetics* / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D003001:Cloning, Molecular; D015870:Gene Expression; D006801:Humans; D008969:Molecular Sequence Data; D010802:Phylogeny; D030162:Reptilian Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D016415:Sequence Alignment; D014714:Vertebrates / Q000145:classification / Q000235:genetics; D055542:alpha-Globins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D055544:beta-Globins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism
chemical_list: D030162:Reptilian Proteins; D055542:alpha-Globins; D055544:beta-Globins