Novel compound participating in lipid metabolism and preparation method thereof

A compound and a new technology, applied in the field of compounds involved in lipid metabolism and their preparation, can solve the problem of low expression

Active Publication Date: 2020-10-23
MEDICINE & BIOENG INST OF CHINESE ACAD OF MEDICAL SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, since most natural product BGCs do not express or express very low levels under laboratory culture conditions, the

Method used

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  • Novel compound participating in lipid metabolism and preparation method thereof
  • Novel compound participating in lipid metabolism and preparation method thereof
  • Novel compound participating in lipid metabolism and preparation method thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0039] Example 1. Overexpression of the key regulatory proteins SGL6295 and SGL6296 in the silent biosynthetic gene cluster Cluster 26

[0040]Using the total genomic DNA of the wild-type strain S. globisporus C-1027 as a template, the regulatory genes SGL6295 and SGL6296 were amplified by PCR, cloned into the pEasy-Blunt Zero vector, and the recombinant plasmid was digested to verify that a band with the same length as the PCR fragment was obtained , after the correct sequence was confirmed by sequencing, it was double digested with NdeI and BamHI and cloned into the pSET152-based expression plasmid pL646 [Hong B, Phornphisutthimas S, Tilley E, Baumberg S, McDowall KJ. Streptomycin production by Streptomyces griseus can be modulated by a mechanism not associated with change in the adpA component of the A-factor cascade. Biotechnol Lett 2007, 29:57-64] NdeI and BamHI sites to obtain recombinant plasmid pL-C26-SGL6295-6296.

[0041] The recombinant plasmid pL-C26-SGL6295-6296 w...

Embodiment 2

[0042] Example 2, Analysis of fermentation products of strains overexpressing regulatory proteins SGL6295 and SGL6296

[0043] The strains overexpressing regulatory proteins SGL6295 and SGL6296 were cultured in liquid fermentation for 7 days, and the fat-soluble secondary metabolites (ethyl acetate extract) were analyzed by TLC and HPLC. TLC results showed that the overexpressed strain C-1027 / pL-C26-SGL6295-6296 had new secondary metabolites compared with the control strain C-1027 / pSET152 ( figure 1 ), and at least seven more color bands than the control strain; HPLC results ( figure 2 ) showed that the overexpression strain had more significant difference peaks at 7-19min, and the compound yield at 30.6min was increased. It shows that after the overexpression of regulatory proteins SGL6295 and 6296, the types and quantities of metabolites increase. Carry out HPLC analysis after scraping the difference band on the TLC plate, show that this band is the product of 30.6min. ...

Embodiment 3

[0044] Example 3, Identification of strain C-1027 / pL-C26-SGL6295-6296 product

[0045] The following method was used to pre-treat and separate the product of strain C-1027 / pL-C26-SGL6295-6296, and after purification by preparative thin-layer chromatography and preparative high-pressure liquid chromatography, the Compounds 1-5 were isolated from SGL6295-6296.

[0046] Extraction and separation of compound 1-5:

[0047] 20L of the fermentation supernatant was extracted with an equal volume of ethyl acetate to obtain 1.1g of the extract. The extracts were mixed with equal amounts of silica gel, and separated by silica gel column chromatography, using chloroform:methanol [100:0 (0.2g, Fr.1), 50:1 (0.1g, Fr.2), 10:1 in sequence (0.12g, Fr.3), 5:1 (0.15g, Fr.4) and 1:1 (0.12g, Fr.5)] mobile phase elution to obtain five components. The above five components were analyzed sequentially by TLC, and the developer was developed using chloroform:methanol:acetic acid 100:20:1, and the ta...

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Abstract

The invention provides a novel compound, the compound is E-13-(2-methylphenyl)-12-ene-tridecanoic acid, and the compound is derived from streptomyces globiformis C-1027, can influence uptake on low density lipoprotein by cells, and participates in an intracellular lipid metabolism process. The compound is white amorphous powder, is insoluble in water, and can be easily dissolved in acetone, chloroform and other low-polarity organic solvents. The invention also provides a method for preparing the compound by using the regulatory protein SGL6295 and the regulatory protein SGL6296.

Description

technical field [0001] The present invention relates to the field of biotechnology, in particular to a compound and its preparation method and application, in particular to a novel compound involved in lipid metabolism and its preparation method. Background technique [0002] Microbial secondary metabolites are an important source of drug discovery. The secondary metabolites produced by Streptomyces have a wide range of biological activities, and more than two-thirds of clinically useful antibiotics are isolated from Streptomyces. Traditional natural product discovery methods have disadvantages such as high compound discovery rate and low discovery rate of new structure natural products. With the reduction of gene sequencing costs and the development of bioinformatics technology, scientists have discovered that there are still a large number of unknown biosynthetic gene clusters (biosynthetic gene clusters, BGCs) in microorganisms, which have great potential for development....

Claims

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Application Information

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IPC IPC(8): C07C57/42C07C51/42C07K14/36C12N15/31C12N15/76C12N1/21C12P7/64C12R1/465
CPCC07C57/42C07C51/42C07K14/36C12P7/6409C12N15/76
Inventor 洪斌江志波李星星任卫聪武临专杜郁胡笑文侍媛媛王丽非张秀敏
Owner MEDICINE & BIOENG INST OF CHINESE ACAD OF MEDICAL SCI
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