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Analysis Interactions of Glycosyltransferase and Substrates by Microscale Thermophoresis (MST) (CAT#: STEM-MB-2446-LGZ)

Introduction

Identification and characterization of key enzymes involved in cell wall biosynthesis and modification is fundamental to gaining insight into cell wall dynamics. However, the very low throughput of glycosyltransferase activity assays, often without acceptor substrates, presents a challenge. Microscale Thermophoresis (MST) enables high-throughput screening of glycosyltransferase substrates.




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.