Preparation method of chlorotoxin

A technology for chlorotoxin and peptide segment, which is applied in the field of preparation of chlorotoxin, can solve problems such as low-cost and large-scale industrial production of chlorotoxin, and achieve the effects of shortening synthesis cycle, reducing side reactions and easy operation.

Active Publication Date: 2022-03-25
杭州禾泰健宇生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

At present, the mass production of chlorotoxin is usually carried out by biological fermentation,

Method used

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  • Preparation method of chlorotoxin
  • Preparation method of chlorotoxin
  • Preparation method of chlorotoxin

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preparation example Construction

[0049] The present invention provides a preparation method of chlorotoxin, which can be achieved by those skilled in the art by referring to the content of this article and appropriately improving the process parameters. In particular, it should be pointed out that all similar replacements and modifications are obvious to those skilled in the art, and they are all considered to be included in the present invention. The method and application of the present invention have been described through preferred embodiments, and relevant personnel can obviously make changes or appropriate changes and combinations to the method and application herein without departing from the content, spirit and scope of the present invention to realize and apply the present invention Invent technology.

[0050] The preparation method of chlorotoxin provided by the invention comprises:

[0051] Step 1: Using Fmoc-AA-OH as raw material, solid-phase step-by-step synthesis to obtain the resin peptide Fmo...

Embodiment 1

[0086] Example 1: Fragment Coupling Method

[0087] step 1:

[0088] Fmoc-Arg(Pbf) 14 -Lys(Boc) 15 -Cys(Trt) 16 -Asp(tBu) 17 -Asp(tBu) 18 -Cys(Trt) 19 -Cys(Trt) 20 -Gly 21 -Gly 22 -Lys(Boc) 23 -Gly 24 -Arg(Pbf) 25 -Gly 26 -Lys(Boc) 27 -Cys(Trt) 28 -Tyr(tBu) 29 -Gly 30 -Pro 31 -Gln(Trt) 32 -Cys(Trt) 33 -Leu 34 -Cys(Trt) 35 -Arg(Pbf) 36 - Synthesis of AM Resin:

[0089] Removal of Fmoc protecting group: Weigh AM Resin (31.646g, 10.0mmol) with a substitution degree of 0.316mmol / g, add it to a solid-phase reaction column, swell the resin with DMF (310ml) for 30 minutes, and drain the solvent. Use DBLK (20% piperidine / DMF, 200ml) to remove the Fmoc protection for 10 minutes, remove the protection twice, and then wash 6 times with DMF (300ml) (take a little resin and add 0.3ml of 5% ninhydrin ethanol solution, Heating at 100 degrees Celsius for 5-10 minutes, the resin will not turn blue).

[0090] Fmoc-Arg(Pbf)-OH coupling: Weigh 12.981g of Fmoc-Arg(Pbf)-OH ...

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Abstract

The invention relates to the technical field of polypeptide synthesis, in particular to a preparation method of chlorotoxin. According to the method, chlorotoxin is prepared through a process of combining solid-phase fragment synthesis and liquid-phase synthesis, and the chlorotoxin is divided into four fragments of 1-5, 6-9, 10-13 and 14-36 for segmented coupling. The method is simple and convenient to operate, and the synthesis period of the process is greatly shortened. Through the solid-liquid combined synthesis method, the yield and purity of chlorotoxin are improved, side reactions and types of by-products are reduced, the total yield can reach 25% or above, and the purity reaches 99% or above.

Description

technical field [0001] The invention relates to the technical field of polypeptide synthesis, in particular to a preparation method of chlorotoxin. Background technique [0002] Chlorotoxin is a peptide component from the venom of the scorpion Leiurius quinquestriatus, the amino acid sequence of which is: [0003] met 1 -Cys 2 -Met 3 -Pro 4 -Cys 5 -Phe 6 -Thr 7 -Thr 8 -Asp 9 -His 10 -Gln 11 -Met 12 -Ala 13 -Arg 14 -Lys 15 -Cys 16 -Asp 17 -Asp 18 -Cys 19 -Cys 20 -Gly 21 -Gly 22 -Lys 23 -Gly 24 -Arg 25 -Gly 26 -Lys 27 -Cys 28 -Tyr 29 -Gly 30 -Pro 31 -Gln 32 -Cys 33 -Leu 34 -Cys 35 -Arg 36 -NH 2 (Disulfide bridge:Cys 2 -Cys 19 ,Cys 5 -Cys 28 ,Cys 16 -Cys 33 ,Cys 20 -Cys 35 ). [0004] Its molecular structural formula is: [0005] [0006] Studies have shown that chlorotoxin can specifically bind to tumor cells, therefore, chlorotoxin can be used as a targeting agent to deliver cytotoxic and / or imaging agents to various tumors, in...

Claims

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Application Information

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IPC IPC(8): C07K14/435C07K1/20C07K1/06C07K1/04
CPCC07K14/43522Y02P20/55
Inventor 钟国庆陈俊华刘艳阳年贺凤高剑
Owner 杭州禾泰健宇生物科技有限公司
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