Derivant with reducing calcium gene related peptide and preparation process thereof

A derivative, calcitonin technology, applied in the field of genetic engineering, can solve the problems of limiting the application value of CGRP, short action time, and high production cost, and achieve the effects of improving short action time, lowering blood pressure, and enhancing myocardial contractility.

Inactive Publication Date: 2005-01-05
FUDAN UNIV
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  • Abstract
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  • Claims
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Problems solved by technology

However, CGRP itself is a small peptide, which is easily degraded in animals, has short action time,

Method used

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  • Derivant with reducing calcium gene related peptide and preparation process thereof
  • Derivant with reducing calcium gene related peptide and preparation process thereof
  • Derivant with reducing calcium gene related peptide and preparation process thereof

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[0026] According to claim 1, the chemically synthesized CGRP gene encodes a total of 37 amino acids, and then the gene is inserted into the E. coli plasmid vector pGEX-4T-2. This recombinant plasmid is named pGEX-CGRP. After subcloning, this recombinant plasmid was then transferred to the E. coli recipient strain TG1. Using double enzyme digestion, PCR amplification and nucleotide sequence analysis, it was confirmed that the sequence of the transformant was consistent with the designed sequence. The transformant can be cultured on LB medium. Pick a single colony, culture it overnight at 37°C, inoculate 1L liquid LB containing 100μg / mL ampicillin at 1:50, culture at 25°C for 5h, add IPTG to the final concentration 0.1mmol / L, induced at 25℃ for 9h. The cells were collected by centrifugation, PBS was added at 1:6, sonicated, and the supernatant was collected after centrifugation. Add Triton-100 to the supernatant to a final concentration of 1%, mix well, then add a certain amount of ...

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Abstract

The invention is a calcitonin gene-related polypeptide (CGRP) derivative and its preparing method. CGRP has important physiologic functions of reducing blood pressure, etc, but is easy to degrade in animal bodies and has short acting time. It designs a segment of DNA sequence linked with the GST genes ofEscherichia coli particle carrier pGEX-4T-2, composing a new expression plasmid pGEX-CGRP with CGRP. It makes separation and extraction on the fusion protein produced by Escherichia coli fermentation to prepare CGRP derivative I. In addition, It clones the genes of the two decompressing small peptides: HHL and PRLKPW to CGRP genes, composing two expressing particles: pGEX-CGRP-3AA and pGEX-CGRP-6AA, transferred into the Escherichia coli cells, then fermenting and separating CGRP derivatives II and III from Escherichia coli cells. The above derivatives can be used in preparing the medicines of curing cardiovascular disease.

Description

technical field [0001] The invention belongs to the technical field of genetic engineering, and specifically relates to a calcitonin gene-related peptide derivative and a preparation method thereof. Background technique [0002] Calcitonin gene-related peptide (CGRP) is widely distributed in the human central nervous system, peripheral nervous system, cardiovascular system, urinary system and other tissues and organs. It has various physiological functions in animals. It is a powerful vasodilator that can dilate blood vessels, lower blood pressure, enhance myocardial contractility, and increase blood output. It has great clinical application value in treating hypertension, regulating arrhythmia, and treating cardiovascular diseases such as heart failure, angina pectoris, coronary heart disease and myocardial necrosis. However, CGRP itself is a small peptide, which is easily degraded in animals, has short action time, high production cost, and high price, which limits the ap...

Claims

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Application Information

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IPC IPC(8): A61K38/23A61P9/04A61P9/06A61P9/10A61P9/12C07K14/585C12N15/12
CPCY02A50/30
Inventor 郑兆鑫严维耀刘明秋邱媛媛
Owner FUDAN UNIV
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