Looking for breakthrough ideas for innovation challenges? Try Patsnap Eureka!

Styrene cyclooxygenase derived from rhizobium, and function of styrene cyclooxygenase

A cyclooxygenase, styrene technology, applied in the direction of oxidoreductase, enzymes, microorganism-based methods, etc., to achieve the effect of excellent enantioselectivity

Active Publication Date: 2020-09-15
CHENGDU INST OF BIOLOGY CHINESE ACAD OF S
View PDF5 Cites 2 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

So far, no styrene cyclooxygenase from rhizobia has been reported

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • Styrene cyclooxygenase derived from rhizobium, and function of styrene cyclooxygenase
  • Styrene cyclooxygenase derived from rhizobium, and function of styrene cyclooxygenase
  • Styrene cyclooxygenase derived from rhizobium, and function of styrene cyclooxygenase

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0021] Example 1 BrSMO heterologous expression

[0022] After the plasmid pET-BrSMO was transformed into Escherichia coli BL21(DE3) competent, a single clone was picked and inoculated in LB medium containing kanamycin (50 mg / L), and cultivated overnight at 37°C and 180 rpm for use as seed liquid. Transfer the seed solution to 200mL TB medium containing kanamycin (50mg / L) with 1% inoculation rate, and cultivate it at 37°C and 180rpm for 3h, when the OD 600 After reaching 0.8, add IPTG (final concentration 0.05mM) and induce at 20°C for 20 hours. Then centrifuge at 8000 rpm at 4° C. for 10 minutes to obtain the bacterial cells, and then resuspend and wash twice with 0.9% NaCl solution to obtain wet bacterial cells. The wet bacteria are used as a biocatalyst for subsequent biocatalysis.

[0023] The obtained wet bacteria were resuspended in potassium phosphate buffer (0.1M, pH 7.0), and the cells were crushed with a high-pressure homogenizer to obtain a cell disruption liquid,...

Embodiment 2

[0024] Example 2 Optimization of styrene cyclooxygenase BrSMO whole cell biocatalysis conditions

[0025] 2.1 Optimum reaction pH value optimization

[0026] The reaction system was 5 mL, including 0.5 g of E. coli (pET-BrSMO) wet cells, 0.1 M potassium phosphate buffer PBK (6.0, 6.5, 7.0, 7.5, 8.0) of different pH, cyclohexane (10%) and Styrene (8mM), shaking reaction at 30°C and 200rpm for 2h. After the reaction was completed, an equal volume of ethyl acetate was used to stop the reaction and extracted, and an appropriate amount of anhydrous sodium sulfate was added to dry it, and the mixture was detected and analyzed by GC. The instrument used is Agilent Technologies 7890B GC gas chromatograph, the chiral column is Cyclodex-B column (30m×0.25mm×0.25μm, USA), the injector temperature, detector sample temperature and column temperature are 260°C and 280°C respectively and 100°C. The results are shown in Table 2. The epoxy production rate is the highest in the 0.1M potassiu...

Embodiment 3

[0035] Embodiment 3 Styrene cyclooxygenase BrSMO carries out biotransformation to different substrates

[0036] The reaction system was 5mL, including 0.1M potassium phosphate buffer (pH 7.0), 0.5g E.coli (pET-BrSMO) wet cells, 100μL isopropanol, 5mM substrate (see Table 4, Table 5), at 30 ℃, 200rpm shaking reaction for 2 hours, stop the reaction with ethyl acetate and extract, add anhydrous sodium sulfate for drying, rotary evaporation to remove the solvent, GC and HPLC detection and analysis, see Table 6 and Table 7. Instrument used for GC detection: Agilent Technologies 7890B GC system-FID detector. Instrument used for HPLC detection: Shimadzu Prominence LC-20AD system-PDA detector.

[0037] It can be seen from Table 4 that the styrene cyclooxygenase BrSMO disclosed in the present invention converts 8 kinds of styrene substrates into corresponding epoxy products, all of which have S-type selectivity. Except for 2a (97% ee) and 5a (95% ee), it showed excellent enantioselec...

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

PUM

No PUM Login to View More

Abstract

The invention discloses a novel styrene mono-cyclooxygenase BrSMO derived from Bradyrhizobium sp. ORS 375, and an application of the novel styrene mono-cyclooxygenase BrSMO in the catalysis of asymmetric oxidation reactions. The enzyme can catalyze styrene or thioether substrates to carry out asymmetric oxidation reactions, and has relatively high activity, wherein the styrene substrates are asymmetrically oxidized to generate S-type epoxy, the enantioselectivity is excellent, and the ee is greater than 99%; and the thioether substrates are asymmetrically oxidized to generate R-type sulfoxide,and the ee value of generated (R)-p-bromophenyl methyl sulfoxide is up to 90%.

Description

technical field [0001] The invention relates to a novel styrene cyclooxygenase BrSMO gene and its catalytic function from Bradyrhizobium sp. engineering field. Background technique [0002] Chirally pure epoxides are important synthetic building blocks for the production of pharmaceuticals and fine chemicals, yet obtaining epoxides with high yields and excellent optical purity remains a great challenge in organic synthesis. To date, many chemical and enzymatic strategies have been developed to synthesize chiral epoxides. Among them, the biocatalytic epoxidation of alkenes is considered to be a green and efficient route to prepare chiral pure epoxy compounds. [0003] Styrene cyclooxygenase, which can catalyze the epoxidation of alkenes to obtain epoxides with excellent enantioselectivity, is an excellent biocatalyst for the preparation of chiral epoxides. At present, many styrene cyclooxygenase genes have been identified from styrene-degrading strains, metagenomes, or thr...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

Application Information

Patent Timeline
no application Login to View More
IPC IPC(8): C12N9/02C12N15/70C12P17/02C12P17/16C12P11/00C12R1/19
CPCC12N9/0071C12Y114/14011C12Y114/14001C12N15/70C12P17/02C12P17/162C12P11/00Y02P20/584
Inventor 吴中柳崔璨郭超刘艳
Owner CHENGDU INST OF BIOLOGY CHINESE ACAD OF S
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Patsnap Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Patsnap Eureka Blog
Learn More
PatSnap group products

Browse by: Latest US Patents, China's latest patents, Technical Efficacy Thesaurus, Application Domain, Technology Topic, Popular Technical Reports.

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap|About US| Contact US: help@patsnap.com