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Analysis Interactions of Bacteriophage Mu DNA Modification Protein Mom with Ligands by Microscale Thermophoresis (MST) (CAT#: STEM-MB-2466-LGZ)

Introduction

Epigenetic modifications play diverse roles in biological systems. Nucleic acid modifications control gene expression, protein synthesis, and sensitivity to nucleolytic enzymes. However, the biosynthetic mechanisms of nucleic acid modification can be challenging. Studying protein-ligand interactions helps decipher the regulatory pathways behind biosynthesis and biological responses. Microscale Thermophoresis (MST) can be used to reveal the biomolecular interactions between the phage Mu DNA modifying protein Mom and its ligands. Compared with traditional methods for studying protein-biomolecule interactions, MST requires less sample size, volume, and analysis time, and thus can screen a large number of candidate molecules that interact with proteins of interest. Another notable feature of this method is that it avoids the need for protein purification, which is often a time- and resource-consuming step, and works well with fully or partially purified cell extracts.




Principle

One of the interacting molecules (mostly proteins) is labeled with fluorescent dye or combined with GFP label. The labeled protein and ligand molecules are placed in the capillary according to a specific concentration gradient. Infrared laser heating generates a microscopic temperature gradient field to undergo thermophoresis, with which the hydration layer, molecular size, electric charge and other molecular properties will change. Then the fluorescence distribution in the reaction system changes. In addition to accurately detecting interactions between biomolecules, MST can also obtain other parameters related to interactions between biomolecules by calculating dissociation constants (Ks), and achieve accurate qualitative analysis.

Applications

For characterizing thermodynamic parameters of biomolecular interactions.

Procedure

1. Sample processing.
2. MST detection.
3. Data analysis.

Materials

• Sample Type: protein 10uM/ 100ul or 50 micrograms, small molecules 100uM/200ul, protein peptides need to be sent at low temperature

Notes

In order to ensure the reliability of the test, some samples should be provided as much as possible.
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