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Complete set of reagents for detecting interaction between protein subjected to post-translational modification and ligand thereof

A post-translational modification and protein technology, which is applied in the field of complete sets of reagents for detecting the interaction between post-translationally modified proteins and their ligands, and can solve the problems of difficult detection of weak protein interactions.

Active Publication Date: 2019-06-21
抟相医药(杭州)有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, weak protein-protein interactions are still difficult to detect

Method used

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  • Complete set of reagents for detecting interaction between protein subjected to post-translational modification and ligand thereof
  • Complete set of reagents for detecting interaction between protein subjected to post-translational modification and ligand thereof
  • Complete set of reagents for detecting interaction between protein subjected to post-translational modification and ligand thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0170] Example 1, Verification of the interaction between H3K9me3 and CD

[0171] In this example, X L is H3K9me3, X is CD, and H3K9me3 interacts with CD.

[0172] 1. Preparation of recombinant vector

[0173] 1. A recombinant vector expressing the fusion protein of SGS and CD

[0174] The DNA fragment (comprising the recognition sequence of NcoI and XhoI) between the NcoI and XhoI recognition sequences of the pRSFDuet-1 vector (Novagen product of Merck) is replaced with the DNA molecule shown in the 12th-1360th position of sequence 2 in the sequence listing, The recombinant vector pRSFDuet-1-SGS was obtained, and pRSFDuet-1-SGS could express the protein shown in Sequence 1 (SGS was fused with His-tag, ie R monomer, denoted as His-SGS).

[0175] Among them, the DNA molecule encoding His-SGS shown in the 14th-1354th position of the sequence 2 shown in the sequence 1, the 1344-1349th and 1355-1360th positions of the sequence 2 are the recognition sequences of NcoI and XhoI re...

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Abstract

The invention discloses a complete set of reagents for detecting interaction between protein subjected to post-translational modification and ligands thereof. The complete set of reagents consists offour reagents A, B, C and D, wherein A is formed by connecting a biomolecule named as R and a protein named as X; B contains a biomolecule named as L; R and L are the same or different and have interaction, and phase change occurs after the R and L interact; C is a polymer formed by a monomer C, the monomer C is a molecule obtained by connecting a monomer named as mc, a reporter group named as I and a biomolecule named as YC, and two or more mc can form the polymer; D is formed by connecting a modified protein named as XL and a biomolecule named as YD; and the YC and the YD have interaction. According to the invention, high enrichment of the interaction protein and the ligand in the phase-change liquid drop is realized, and the weak interaction signal is amplified, so that the detection iseasy.

Description

technical field [0001] The invention relates to a set of reagents for detecting the interaction between a post-translation modified protein and its ligand in the field of biotechnology. Background technique [0002] As a characteristic of matter, "phase transition" has long been known in the physics world and in daily life. In recent years, scientists have gradually discovered that phase transition (or phase separation) mechanisms also widely exist in biological cells, and in the time and space of the cell life cycle Regulation and other aspects to perform important biological functions. [0003] Current research has found that when multivalent macromolecules in solution interact with their multivalent ligands, larger complexes tend to be produced, and the solubility of the latter generally decreases, thereby separating from the common solution phase to form a complex This transition process is called "liquid-liquid separation phase transition". Wherein, the multivalent va...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/53G01N33/68
CPCG01N33/566G01N33/573
Inventor 李丕龙李剑王静周敏李维平王亮吴荣波陈鹤
Owner 抟相医药(杭州)有限公司
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