Enzymatic synthesis method of S-alpha-D-glucoside-L-cysteine

A technology of cysteine ​​and glucoside, applied in the field of enzymatic synthesis of S-α-D-glucoside-L-cysteine, to achieve the effect of improving substrate utilization

Inactive Publication Date: 2012-12-19
NANJING TECH UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0009] The technical problem to be solved by the present invention is to provide an enzymatic synthesis method of S-α-D-glucoside-L-cysteine ​​to overcome the defects of the chemical synthesis method

Method used

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  • Enzymatic synthesis method of S-alpha-D-glucoside-L-cysteine
  • Enzymatic synthesis method of S-alpha-D-glucoside-L-cysteine
  • Enzymatic synthesis method of S-alpha-D-glucoside-L-cysteine

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0038] Embodiment 1 prepares and purifies sucrose phosphorylase

[0039] Bacteria: Leuconostoc mesenteroides ATCC12291

[0040] Seed activation medium: 10g / L peptone, 10g / L meat extract, 5g / L yeast extract, 20g / L glucose, 5g / L sodium acetate trihydrate crystals, 1ml / L Tween 80, 2g / L citric acid Triammonium, 2g / L K 2 HPO 4 , 20g / L agar, 0.2g / LMgSO 4 ·7H 2O, 0.05g / L MnSO 4 2H 2 O, pH5.4.

[0041] Fermentation medium: 20g / L sucrose, 10g / L soybean peptone, 10g / L yeast extract, 10g / LK 2 HPO 4 , 10ml / L Vitamin-mineral stock solution (1L contains VB 1 1g; VC 5g; anhydrous magnesium sulfate 40g; manganese sulfate monohydrate 30g; ferrous sulfate heptahydrate 1g), pH 7.6.

[0042] Sterilization conditions: Separately sterilize the glucose and metal ions in the seed activation medium; separately sterilize the sucrose in the fermentation medium, sterilize the Vitamin-mineral stock solution with a sterile filter, and sterilize the others by autoclaving at 121°C for 15 minutes. ...

Embodiment 2

[0048] Get 200 U of the sucrose phosphorylase obtained in Example 1, add 0.4 mol / L sucrose, 0.1 mol / L L-cysteine, 10 ml of MES buffer (pH 5.5, 10 mM), mix well and then stir The reaction was carried out for 13 hours, and the reaction temperature was 30°C. After the reaction, the enzyme was removed by microfiltration, and detected by Dionne U3000 detection system, and the conversion rate of L-cysteine ​​was 5.2%.

Embodiment 3

[0050] Get 200U of the sucrose phosphorylase obtained in Example 1, add to 0.5mol / L sucrose, 0.15mol / L L-cysteine, 10ml of MES buffer (pH7.0, 10mM), mix well and then stir The reaction was carried out for 12 hours, and the reaction temperature was 35°C. After the reaction, the enzyme was removed by microfiltration, and detected by Dionne U3000 detection system, and the conversion rate of L-cysteine ​​was 10.1%. The liquid chromatogram is as figure 1 , take a sample to make a mass spectrum, the mass spectrum is as follows figure 2 shown.

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Abstract

The invention discloses an enzymatic synthesis method of S-alpha-D-glucoside-L-cysteine. Aqueous solution containing sucrose and L-cysteine is used as a substrate, sucrose phosphorylase is added, and the S-alpha-D-glucoside-L-cysteine is prepared by transglucose-based reaction of the sucrose phosphorylase. Compared with a chemical synthesis method, group protection and removal are not needed and the steps are simple in the method; the reaction raw materials are simple, easily obtained and environment-friendly; and the sucrose phosphorylase is obtained by simple fermentation of leuconostosm esenteroides, so the cost is low.

Description

technical field [0001] The invention belongs to the field of biochemical industry and relates to an enzymatic synthesis method of S-α-D-glucoside-L-cysteine. Background technique [0002] Oligosaccharide complexes play an important role in the biological activities of various organs of living tissues, such as: cell recognition, information transmission between cells and cell adhesion process. The carbohydrate group attached to the polypeptide can increase its water solubility, and can also be used as a targeted drug to target biologically active compounds. The glucosinolate formed by connecting the cysteine-SH group with a glucose group has high chemical stability and has great application in the synthesis of polypeptide glucosinolate derivatives. The application of glucosinolates in anticancer drugs has been further expanded in recent years, leading to the great application of cysteine ​​glucosinolates in the synthesis of anticancer drugs and targeted drugs in the process ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12P19/64
Inventor 姜岷侯顾伟马江锋隋姗姗奚永兰陈可泉韦萍欧阳平凯
Owner NANJING TECH UNIV
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