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Bacterial Expression of Rhamnose-Binding Lectin from Catfish Eggs
Shigeki Sugawara1, Takeo Tatsuta1, Masahiro Hosono2
1Division of Cell Recognition Study, Institute of Molecular Biomembrane and Glycobiology, Tohoku Medical and Pharmaceutical University, Sendai, Japan.
Methods in Molecular Biology (Clifton, N.J.)
|April 20, 2020
Summary
Researchers found that specific conditions can improve the solubility of Silurus asotus lectin (SAL) recombinants. Highly basic environments and the presence of dithiothreitol (DTT) help keep these rhamnose-binding lectin (RBL) proteins soluble when expressed in E. coli.
Area of Science:
- Biochemistry
- Molecular Biology
- Structural Biology
Background:
- SUEL-like lectins, or rhamnose-binding lectins (RBLs), possess unique tandemly repeated carbohydrate-recognition domains.
- These lectins exhibit α-galactoside-binding capabilities, distinguishing them within animal lectin families.
- RBLs are often found in inclusion bodies when expressed in Escherichia coli.
Purpose of the Study:
- To describe methods for the expression and refolding of Silurus asotus lectin (SAL).
- To identify conditions that promote the solubility of SAL recombinants.
- To utilize E. coli KRX as a host strain for SAL expression.
Main Methods:
- Expression of Silurus asotus lectin (SAL) in E. coli KRX host strain.
- Refolding protocols for recombinant SAL.
- Investigating the impact of solution conditions (pH, reducing agents) on protein solubility.
Main Results:
- Highly basic conditions were found to be beneficial for SAL solubility.
- The addition of dithiothreitol (DTT) significantly enhanced the solubility of SAL recombinants.
- Optimal conditions were identified for maintaining soluble SAL protein.
Conclusions:
- Basic pH and reducing conditions are crucial for soluble expression of SAL.
- These findings provide a method for producing functional RBLs in E. coli.
- The study contributes to understanding the expression and refolding of a specific α-galactoside-binding lectin.
