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The Physiological Activities Study of Insulinotropic Peptides from Three Amphibian Skins

Author: LiZongJie
Tutor: LaiZuo
School: Nanjing Agricultural College
Course: Zoology
Keywords: Amphibians Insulinotropic peptide Skin secretion Insulin secretion Insulin release
CLC: R285.5
Type: Master's thesis
Year: 2008
Downloads: 32
Quote: 0
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Abstract

Insulinotropic peptide is a bioactive peptides can regulate insulin secretion, and to study the structural characteristics, physiological activity, as well as clinical drug application for diabetes II opened up new therapeutic approach and treatment philosophy. Peptide toxin as a molecular probe, is conducive to the further insight into the mechanisms of insulin secretion signal conditioning and understanding of the pathogenic mechanism of certain toxins. At present, the international community about the pharmacological effects of glucagon-like peptide -1 (GLP-1) and its analogues basic research and clinical drug development progress very rapidly, and foreign scholars from insects, amphibians, reptiles and other animals found in the insulinotropic peptide. However, the very few scholars engaged natural insulinotropic releasing peptide Isolation and identification of research work. The natural advantage of broad multi-species biological resource library for our research, isolated and purified insulinotropic peptide and explore the mechanism can lay an important foundation for the development of this field of research platforms and fill the blank of domestic research The study has important and far-reaching significance. In this study, no means the disk Odorrana (Odorrana grahami), the brown point Amolops (Amolops loloensis) and Black Belt frog (Rana nigrovittata), three distribution in Yunnan amphibians as experimental material, mainly through Sephadex ( Sephadex G-50) molecular sieve filter and means of reversed-phase high performance liquid chromatography (RP-HPLC) separation and purification of frog skin secretion, insulin releasing activity over ten separation peak. Eight insulinotropic peptide amino acid sequence separated peaks were measured relative molecular mass use of flight mass spectrometry, and Edman degradation, were named as: odorranain-G (thirteen peptide), odorranain-Z (20 peptides) , odorranain-L1 (peptide) twenty-one, odorranain-Y (11 peptides) odorranain-X1 (sixteen peptide), odorranain-X2 (sixteen peptide) amolopin (16 peptides) and nigroain-A (ten heptapeptide);, and mRNA was extracted using living frog, the cDNA library was constructed according to the known conserved signal peptide sequence primers were designed, and the PCR amplification product was transformed into E. coli and sequenced positive clones, and the amino acid sequence and the separation and purification of gene cloning by the polypeptide translation obtained The polypeptide obtained by N-terminal sequencing results are consistent. Rat islets and INS-1 pancreatic β cell lines were cultured in vitro detection of two model insulinotropic activity, ^{125 I radioactive immunoassay detects insulin levels. The calcium fluorescence probe Fluo-3 and laser confocal microscopy study confirmed: promoting molecular mechanism of beta cells to secrete insulin-releasing peptide stimulation is not due to increased calcium influx caused changes in intracellular calcium ion concentration. Because diabetes can cause a variety of complications of serious harm, the insulinotropic peptide activity is necessary. This study shows that, The insulinotropic peptide odorranain-G also has anti-bacterial, fungal activity, seen the polypeptide in hypoglycemic while also can control diabetes caused by various infections, there is a good prospect of application. Odorranain-Z angiogenesis inhibitory activity, the current study the treatment of diabetic retinopathy hot issues, so odorranain-Z also has a very good potential for drug development. In this study, the first time in the country established the insulinotropic peptide separation and purification, structural analysis and activity assay technology research platform, and successfully obtained a structure of eight insulinotropic peptide, are reported insulinotropic peptide amino acid sequences are significantly different, that are a new biologically active peptide. Which never refers to a large number of the insulinotropic peptide skin a disk Odorrana (Odorranagrahami), suggesting that the animal skin secretion is a valuable biological resource library, and odorranain-Y by only eleven amino acid residues, has been found to promote insulin-releasing peptide molecular weight minimum. Promote the physiological activity of the insulin-releasing peptide research provides a new research direction and broad prospects for improving insulin secretory function of patients with type II diabetes.}

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