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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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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

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 application value of CGRP in medical practice.

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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Embodiment Construction

[0026] According to claim 1, the chemically synthesized CGRP gene encodes a total of 37 amino acids, and then inserts the gene into the Escherichia coli plasmid vector pGEX-4T-2, and this recombinant plasmid is named pGEX-CGRP. After subcloning, the recombinant plasmid was transformed into Escherichia coli recipient strain TG1. Double digestion, PCR amplification and nucleotide sequence analysis were used to confirm that the sequence of the transformant was consistent with the designed sequence. Transformants can be cultured on LB medium, pick a single colony, culture overnight at 37°C, inoculate into 1L liquid LB containing 100 μg / mL ampicillin at a ratio of 1:50, culture at 25°C for 5 hours, add IPTG to a final concentration of 0.1mmol / L, induced at 25°C for 9h. Collect the cells by centrifugation, add PBS at a ratio of 1:6, sonicate, and collect the supernatant after centrifugation. Add Triton-100 to the supernatant to a final concentration of 1%, mix well, then add a cer...

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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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