title: Full-length cDNA cloning and structural characterization of preproinsulin in Alligator sinensis.
abstract: Insulin is an important endocrine hormone that plays a critical physiological role in regulating metabolism and glucostasis in vertebrates. In this study, the complete cDNA of Alligator sinensis preproinsulin gene was cloned for the first time by reverse transcription-polymerase chain reaction and rapid amplification of cDNA ends methods; the amino acid sequence encoded and protein structure were analyzed. The full-length of preproinsulin cDNA sequence consists of 528 base pairs (bp), comprising a 34-bp 5'-untranslated region, a 170-bp 3'-untranslated region and an open reading frame that is 324 bp in length. The open reading frame encodes a 107-amino acid preproinsulin with a molecular weight of approximately 12,153.8 Da, theoretical isoelectric point of 5.68, aliphatic index of 92.06, and grand average of hydropathicity of -0.157, from which a signal peptide, a B-chain, a C-peptide, and an A-chain are derived. Online analysis suggested that the deduced preproinsulin amino acid sequence contains a transmembrane region, and that it has a signal peptide whose cleavage site occurs between alanine 24 and alanine 25. Comparative analysis of preproinsulin amino acid sequences indicated that the A-chain and B-chain sequences of preproinsulins are highly conserved between reptiles and birds, and that the preproinsulin amino acid sequence of Alligator sinensis shares 89% similarity to that of Chelonia mydas, but low similarity of 48-63% to those of mammals and fishes. The phylogenetic tree constructed using the neighbor-joining method revealed that preproinsulin of Alligator sinensis had high homology with reptiles and birds, such as Chelonia mydas, Gallus gallus, and Columba livia.
authors.affiliation: College of Life Sciences, Anhui Normal University, Wuhu, Anhui, China.
authors.forename: R
authors.initials: R
authors.lastname: Zhang
authors.affiliation: College of Life Sciences, Anhui Normal University, Wuhu, Anhui, China szzhang@mail.ahnu.edu.cn.
authors.forename: S Z
authors.initials: SZ
authors.lastname: Zhang
authors.affiliation: College of Life Sciences, Anhui Normal University, Wuhu, Anhui, China.
authors.forename: E
authors.initials: E
authors.lastname: Li
authors.affiliation: College of Life Sciences, Anhui Normal University, Wuhu, Anhui, China.
authors.forename: C
authors.initials: C
authors.lastname: Wang
authors.affiliation: Alligator Research Center of Anhui Province, Xuanzhou, China.
authors.forename: C L
authors.initials: CL
authors.lastname: Wang
authors.affiliation: College of Life Sciences, Anhui Normal University, Wuhu, Anhui, China.
authors.forename: X B
authors.initials: XB
authors.lastname: Wu
mesh_terms: D000489:Alligators and Crocodiles / Q000235:genetics*; D000595:Amino Acid Sequence; D000818:Animals; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000737:chemistry / Q000235:genetics*; D007328:Insulin / Q000145:classification / Q000235:genetics*; D008969:Molecular Sequence Data; D010802:Phylogeny; D011498:Protein Precursors / Q000145:classification / Q000235:genetics*; D021382:Protein Sorting Signals / Q000235:genetics; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid
chemical_list: D018076:DNA, Complementary; D007328:Insulin; D011498:Protein Precursors; D021382:Protein Sorting Signals; C013737:preproinsulin