Preparation and application of recombinant mannase-glp-1 and its homologues

A technology of GLP-1 and recombinant engineering bacteria, which is applied in the direction of recombinant DNA technology, biochemical equipment and methods, DNA/RNA fragments, etc., can solve the problems such as the limitations of oral administration, and achieve good application value and good reduction The effect of high blood sugar and protein expression

Active Publication Date: 2021-12-31
ANHUI NEW SIMON BIOTECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, both albiglutide and dulaglutide are injected once a week, which cannot achieve the limitations of oral administration

Method used

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  • Preparation and application of recombinant mannase-glp-1 and its homologues
  • Preparation and application of recombinant mannase-glp-1 and its homologues
  • Preparation and application of recombinant mannase-glp-1 and its homologues

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0049] This embodiment provides a high-efficiency expression and purification of mannanase and GLP-1 recombinant fusion protein (MANNase-GLP-1), including the following steps:

[0050] (1) Use primer pairs to clone the target fragment encoding GLP-1 and mannanase, perform double enzyme digestion after PCR amplification, and then connect the obtained gene sequence encoding GLP-1 and mannanase to the pPICZαA plasmid , the recombinant expression vector pPICZαA-MANNase-31P was constructed;

[0051] The gene sequences encoding GLP-1 and mannanase are shown in SEQ ID NO.1 and SEQ ID NO.2 respectively; the sequence of the primer pair for amplifying the gene sequence encoding GLP-1 is shown in SEQ ID Shown in NO.3 and shown in SEQ ID NO.4; The sequence of the gene sequence primer pair for amplifying the gene sequence encoding mannanase is shown in SEQ ID NO.5 and shown in SEQ ID NO.6; the fusion The amino acid sequence of protein MANNase-31P is shown in SEQ ID NO.7;

[0052] (2) Tra...

Embodiment 2

[0062] This embodiment provides a high-efficiency expression and purification of mannanase and GLP-1 recombinant fusion protein, including the following steps:

[0063] (1) Use primer pairs to clone the target fragment encoding GLP-1 and mannanase, perform double enzyme digestion after PCR amplification, and then connect the obtained gene sequence encoding GLP-1 and mannanase to the pPICZαB plasmid , the recombinant expression vector pPICZαB-MANNase-31P was constructed;

[0064] The gene sequences encoding GLP-1 and mannanase are shown in SEQ ID NO.1 and SEQ ID NO.2 respectively; the sequence of the primer pair for amplifying the gene sequence encoding GLP-1 is shown in SEQ ID As shown in NO.3 and shown in SEQ ID NO.4, the sequence of the gene sequence primer pair for amplifying the gene sequence encoding mannanase is shown in SEQ ID NO.5 and shown in SEQ ID NO.6; the fusion The amino acid sequence of protein MANNase-31P is shown in SEQ ID NO.7;

[0065] (2) Transform the re...

Embodiment 3

[0075] This embodiment provides a high-efficiency expression and purification of mannanase and GLP-1 recombinant fusion protein, including the following steps:

[0076] (1) Use primer pairs to clone the target fragment encoding GLP-1 and mannanase, perform double enzyme digestion after PCR amplification, and then connect the obtained gene sequence encoding GLP-1 and mannanase to the pPICZαC plasmid , the recombinant expression vector pPICZαC-MANNase-31P was constructed;

[0077]The gene sequences encoding GLP-1 and mannanase are shown in SEQ ID NO.1 and SEQ ID NO.2 respectively; the sequence of the primer pair for amplifying the gene sequence encoding GLP-1 is shown in SEQ ID Shown in NO.3 and shown in SEQ ID NO.4; The sequence of the gene sequence primer pair for amplifying the gene sequence encoding mannanase is shown in SEQ ID NO.5 and shown in SEQ ID NO.6; the fusion The amino acid sequence of protein MANNase-31P is shown in SEQ ID NO.7;

[0078] (2) Transform the recomb...

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Abstract

The invention relates to the high-efficiency expression, purification and application of mannanase and its homologues and GLP-1 recombinant fusion protein. Through gene recombination technology, the mannanase and its Homologs and GLP-1 recombinant fusion protein, obtained high-yield mannanase and its homologues and GLP-1 recombinant fusion protein through the separation and purification method of filtration and concentration, with high fermentation expression and simple separation steps, It solves the limitations of the current GLP‑1 analogues, such as low yield, high cost, and frequent injections. The mannanase of the present invention and its homologue and GLP-1 recombinant fusion protein not only have the hypoglycemic effect of the injection, but also can lower the blood sugar and reduce body weight through oral administration, and have a great effect on obese and prediabetic patients. Good application value. At the same time, it provides a basis for further research on the mechanism of lowering blood sugar and weight loss of mannanase and its homologues and GLP-1 recombinant fusion protein.

Description

technical field [0001] The invention belongs to the technical field of biomedicine, and specifically relates to the high-efficiency expression, purification and application of mannanase and its homologue and GLP-1 recombinant fusion protein. Background technique [0002] Diabetes is one of the important diseases that seriously threaten the life and health of human beings around the world in modern society. Diabetes mellitus is an endocrine-metabolic syndrome characterized by chronic hyperglycemia. According to the National Health and Family Planning Commission, 50.1% of adults are currently in pre-diabetes, and they may become new diabetics every day. More data shows that the prevalence of diabetes in China has grown rapidly in recent decades: from 0.67% in the 1980s to 11.6% in 2010, an increase of 17 times, and it is increasingly younger. [0003] In recent years, glucagon-like peptide-1 (GLP-1) not only has an excellent hypoglycemic effect, but also has the characterist...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/81C12N15/62C12N15/11C07K19/00A61K38/47A61K38/26A61K47/64A61K47/55A61P3/10A61P3/04
CPCC12N9/2488C12N15/815C07K14/605A61K38/26A61K38/47A61K47/64A61K47/55A61P3/10A61P3/04C12Y302/01025C07K2319/00C12R2001/84C12N1/165
Inventor 姜伟楼慧强王岩余卫雄徐冲
Owner ANHUI NEW SIMON BIOTECH CO LTD
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