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Construction method for double-promoter yeast random peptide library for screening protein dissociation peptide

A random peptide library and dual-promoter technology, applied in the fields of molecular biology and biopharmaceuticals, can solve problems such as unsatisfactory treatment effects, and achieve the effect of saving tedious procedures

Inactive Publication Date: 2006-09-06
INST OF BASIC MEDICAL SCI ACAD OF MILITARY MEDICAL SCI OF PLA
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Problems solved by technology

Another example is that the concentration of free insulin-like growth factor IGF-1 in the body is closely related to diabetes. At present, the treatment of diabetes is mainly based on Chinese and Western medicines, and it is controlled by adjusting diet. The overall therapeutic effect is not ideal.

Method used

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  • Construction method for double-promoter yeast random peptide library for screening protein dissociation peptide
  • Construction method for double-promoter yeast random peptide library for screening protein dissociation peptide
  • Construction method for double-promoter yeast random peptide library for screening protein dissociation peptide

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Embodiment

[0027] Example, Screening of BAFF-R / TARF-C Complex Dissociation Peptides

[0028] BAFF (B-cell-activating factor) is a member of the TNF family discovered in 1999. It plays an important role in the pathogenesis of autoimmunity and malignant B lymphoma by affecting the survival and maturation of B cells. BAFF mainly regulates B cells through BAFF-R, and TARF3, as a downstream molecule of BAFF-R-mediated alternative NF-κB pathway, plays an important role in signal transduction. We transferred the intracellular region of BAFF-R and the TRAF domain (TRAF-C) gene of TRAF3, constructed pBD-BAFF-R and pAD-TRAF-C recombinant expression vectors, and co-transfected yeast Y190 host cells. Hybridization verified the interaction between BAFF-R intracellular region and TRAF-C. And in the second conditional methionine promoter of pBridge (P MET25 ) was inserted downstream of a random 10-peptide library with thioredoxin as the backbone, and the storage capacity and diversity of the random p...

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Abstract

The invention discloses a method for construction of double-activation subyeast random peptide library for screening dissociating peptide of protein, which belongs to the molecular biology and biopharmaceutics categories. Making use of characteristic that the DNA binding domain (BD) and the activation domain (AD) of the yeast activator of transcription can separate in space, the method consists of inserting gene X in domain BD of Bridge particles, inserting random 10 peptide library with thioredoxin as framework in second conditional methilanin promoter (PMET25) downstream and inserting gene Y in pACT2 containing domain AD, co-transforming yeast host cell, finally screening small peptide molecule which can promote X-Y dissociation by using the expression of auxotrophic gene and the corresponding narrative gene. The method provides a new approach for studying dissociation of protein complex; the obtained peptide of protein can effectively block up the important protein complex in the signal path, which provides a target point for medicines which can cure diseases caused by abnormal signal transduction path.

Description

technical field [0001] The invention belongs to the field of molecular biology and biopharmaceutical technology, and particularly relates to a method for constructing a double-promoter yeast random peptide library for screening protein dissociation peptides. Background technique [0002] In life activities, abnormal signal transduction pathways lead to diseases. For example, B cell activating factor BAFF (B-cell-activating factor) mainly affects the survival and maturation of B cells through BAFF-R, and BAFF-R mediated Intracellular signaling pathways play an important role in the pathogenesis of autoimmunity and malignant B lymphoma. The small peptides screened by this method can promote the dissociation of the BAFF-RTARF3 complex, block the BAFF-R signaling pathway, and achieve the purpose of treating autoimmune diseases and malignant B lymphoma. Another example is that the concentration of free insulin-like growth factor IGF-1 in the body is closely related to diabetes. ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/19C12N15/81C12Q1/68A61K38/08A61K38/10
CPCY02A50/30
Inventor 徐东刚杜建芳魏平邹民吉王嘉玺
Owner INST OF BASIC MEDICAL SCI ACAD OF MILITARY MEDICAL SCI OF PLA
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