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Detection of silk protein transitions by Differential Scanning Fluorimetry(DSF) (CAT#: STEM-MB-0754-WXH)

Introduction

Silk is made up of two primary proteins; a fibrous protein known as fibroin, and a sticky protein known as sericin, with the two comprising 70–80% and 20–30% of silk, respectively. Sericin has a high hydroxy amino acid content which is important for the water-binding capacity which regulates the skin's moisture content. It also has a unique carbohydrate moiety and a unique repetitive amino acid sequence which give it a high affinity for bonding to adhering proteins resulting in a tightening, anti-wrinkle effect. In addition, because of its high molecular weight, it leaves a substantive semi-occlusive film that persists even after washing, and can increase the skins permeability.




Principle

Differential Scanning Fluorimetry measures protein thermal unfolding by monitoring changes in fluorescence emission of a sample upon heating. This allows the determination of protein thermostability and complex formation even with weakly binding ligands by thermal shift assay. Differential Scanning Fluorimetry is therefore ideally suited for screening of optimum buffer conditions like pH, buffer composition and ionic strength. The technique is applicable to any biological sample, from soluble proteins to integral membrane proteins.

Applications

To identify low-molecular-weight ligands that bind and stabilize purified proteins.
To measure the denaturation and unfolding of proteins.

Procedure

1. Preparation of compound solutions
2. Preparation of buffer/additive screen plates
3. Preparation of compound storage plates
4. Equipment preparation
5. Sample preparation
8. Performing the scan

Materials

Real-time PCR instrument