Novel method for rapidly removing removable framework modification and application

A new method and removal technology, applied in the field of preparation of polypeptides or proteins, can solve the problems of difficult removal and slow removal, and achieve the effects of reducing aggregation, inhibiting the formation of secondary structures, and high yield

Active Publication Date: 2021-08-17
UNIV OF SCI & TECH OF CHINA
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0005] After research, it is found that the RBM on the polypeptide sequence is slowly removed in peptide cleavage reagents such as TFA, mainly because there is an electron-withdrawing group such as nitro on the benzene ring of RBM. If the nitro group on the RBM is removed, it will be transformed into In order to be extremely sensitive to the Hmb structure of TFA, the RBM on the polypeptide sequence can be easily removed, but it is very difficult to directly remove the nitro group on the RBM in chemically synthesized proteins

Method used

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  • Novel method for rapidly removing removable framework modification and application
  • Novel method for rapidly removing removable framework modification and application
  • Novel method for rapidly removing removable framework modification and application

Examples

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preparation example Construction

[0087] The fourth aspect of the present invention provides a method for preparing a polypeptide, comprising the following steps:

[0088] S1, reacting 2-hydroxy-4-methoxy-5 nitrobenzaldehyde with the amino group at the N-terminal of the polypeptide on the solid-phase resin to generate a carbon-nitrogen double bond structure to obtain the first polypeptide compound. Since it is a prior art, no longer elaborate;

[0089] S2. In an organic solution, reducing the first polypeptide compound with a reducing agent to reduce the carbon-nitrogen double bond structure to a carbon-nitrogen single bond structure to obtain a second polypeptide compound, the purpose of which is to reduce the introduction of RBM carbon-nitrogen double bond, in this step, the organic solution can be a conventional choice, such as DMF solution, the reducing agent here is not particularly limited, can be a conventional choice in the art, specific examples include but are not limited to NaBH 4 , sodium triaceto...

Embodiment 1

[0096] This example discloses a method for preparing a model peptide, comprising the following steps:

[0097] S1, contacting 2-hydroxy-4-methoxy-5 nitrobenzaldehyde with the polypeptide on the solid phase resin shown in formula (a1), to obtain the polypeptide on the solid phase resin shown in formula (a2),

[0098]

[0099] Wherein, the solid-phase resin shown in formula (a1) is obtained by Fmoc solid-phase synthesis method, and the specific steps are: adding Rink AM resin to a polypeptide synthesis tube, and using N,N dimethylformamide (DMF) / dichloromethane ( DCM) (volume ratio 1:1) was swollen for 30 minutes, then 20% piperidine in DMF solution was added to treat the resin for 5 minutes and 10 minutes, and the resin was washed three times with DMF, DCM and DMF successively. Then 3.8eq of 2-(7-azobenzotriazole)-tetramethyluronium hexafluorophosphate (HATU), 8eq of N,N diisopropylethylamine (DIEA) and 4eq of Fmoc-protected The first amino acid at the C-terminal of the tar...

Embodiment 2

[0125] This example adopts the same implementation method as Example 1, except that the reaction temperature in step S52 is 0°C.

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Abstract

The invention discloses a novel method for rapidly removing RBM (Removable Framework Modification) and application, and the novel method for rapidly removing RBM comprises the following steps: firstly reducing nitro-NO2 in RBM introduced into polypeptide into amino -NH2, then converting the amino -NH2 into diazonium salt (nitrogen in triple linkage with nitrogen), and then removing the diazonium salt (nitrogen in triple linkage with nitrogen) by a reducing agent, so as to achieve convertion of RBM into 2-hydroxy-4-methoxyl benzyl (HMB). Therefore, the RBM is rapidly removed from polypeptide through a polypeptide cutting reagent, in addition, the method is easy to operate and high in yield, the method can be applied to preparation of polypeptide or protein, and the synthetic yield of polypeptide can be effectively improved, chemical connection of protein is assisted, and natural polypeptide or protein is obtained.

Description

technical field [0001] The invention belongs to the technical field of polypeptide or protein synthesis, and in particular relates to a conversion method for converting RBM of a removable backbone modification on a polypeptide sequence into HMB, and a new method for rapidly removing a removable backbone modification of a polypeptide by using the conversion method , and the application of the new method for rapid removal of the detachable backbone modification in the preparation of polypeptides or proteins. Background technique [0002] Chemical protein synthesis can obtain proteins with post-translational modifications or site-specific tags, and while this technique is finding increasing application in the study of water-soluble globular proteins, the chemical synthesis of difficult-sequence peptides remains a challenge. huge challenge. Specifically, difficult-sequence peptides are difficult to synthesize mainly because they form secondary structures (α-helices or β-sheets)...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K1/04C07K1/06C07K7/06C07K14/00
CPCC07K1/04C07K1/061C07K7/06C07K14/001Y02P20/55
Inventor 郑基深薛敏李影周永康
Owner UNIV OF SCI & TECH OF CHINA
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