Aggregation Prevention Assay for Chaperone Activity of Proteins Using Spectroflurometry
Manish Bhuwan1, Madhuri Suragani2, Nasreen Z Ehtesham3
1Molecular Infection and Functional Biology Laboratory, Kusuma School of Biological Sciences, Indian Institute of Technology, New Delhi, India.
Bio-Protocol
|August 30, 2021
Summary
Molecular chaperones prevent protein aggregation by binding to nonnative protein conformations. This study details an aggregation prevention assay to evaluate the chaperone activity of resistin and MoxR1 proteins.
Area of Science:
- Biochemistry
- Molecular Biology
- Protein Science
Background:
- Molecular chaperones are essential proteins that stabilize other proteins against thermal aggregation.
- Chaperone activity involves binding to nonnative protein conformations and facilitating proper folding through dynamic association and dissociation cycles.
- The aggregation prevention assay is a key method for assessing chaperone function using heat-labile proteins like Maltodextrin glucosidase (MalZ), Citrate Synthase (CS), and NdeI.
Purpose of the Study:
- To describe a detailed protocol for the aggregation prevention assay.
- To evaluate the chaperone activity of two novel proteins, resistin and MoxR1, in preventing thermal aggregation of model substrate proteins.
Main Methods:
- Utilized the aggregation prevention assay to assess chaperone activity.
- Tested the ability of human resistin (hRes) and Mycobacterium tuberculosis MoxR1 to prevent thermal aggregation of MalZ, CS, and NdeI.
Main Results:
- The study successfully outlines a protocol for the aggregation prevention assay.
- Resistin and MoxR1 were analyzed for their efficacy in preventing the thermal aggregation of MalZ, CS, and NdeI.
Conclusions:
- The aggregation prevention assay provides a definitive method for determining chaperone activity.
- Resistin and MoxR1 demonstrate potential chaperone activity, warranting further investigation into their mechanisms.
