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Protein refolding in reversed micelles: Interactions of the protein with micelle components

Protein refolding in reversed micelles: Interactions of the protein with micelle components 10.1002/bit.260351003.abs A novel process has been developed to improve the refolding yield of denatured proteins. It uses reversed micelles to isolate denatured protein molecules from each other and thus, upon refolding, reduces the intermolecular interactions which lead to aggregation. The feasibility of this process was first demonstrated with Ribonuclease A as a model protein. In the present work, we expanded the scope of this study to better understand both the general mechanisms of protein refolding in reversed micelles and the biotechnological applicability of the process. First, we investigated the interactions between the individual components of the reversed micellar system (the protein molecule, the denaturant guanidine hydrochloride (GuHCl), and the surfactant (AOT)) during the refolding process. We then extended our studies to a more hydrophobic protein, γ‐interferon, which aggregates upon refolding in aqueous solution. However, it was also found to aggregate in our reversed micelle process during the extraction step. Since γ‐interferon is a much more hydrophobic protein than RNase, we hypothesize that interactions between hydrophobic amino acids and the surfactant layer may interfere with refolding. This hypothesis was tested by studying the refolding of chemically modified RNase. The substitution of 55% of the surface lysine residues with hydrophobic caproyl groups caused a significant decrease in the refolding yield of RNase in the reversed micellar system without affecting aqueous solution renaturation. In addition, the extraction efficiency of the enzyme from reversed micelles back into aqueous solution was severely reduced and resulted in aggregation. These experiments indicate that unfolded hydrophobic Proteinsinteract with the Surfactant molecules, which limits their ability to refold in reversed micelles. http://www.deepdyve.com/assets/images/DeepDyve-Logo-lg.png Biotechnology and Bioengineering Wiley

Protein refolding in reversed micelles: Interactions of the protein with micelle components

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References (28)

Publisher
Wiley
Copyright
Copyright © 1990 John Wiley & Sons, Inc.
ISSN
0006-3592
eISSN
1097-0290
DOI
10.1002/bit.260351003
pmid
18588241
Publisher site
See Article on Publisher Site

Abstract

10.1002/bit.260351003.abs A novel process has been developed to improve the refolding yield of denatured proteins. It uses reversed micelles to isolate denatured protein molecules from each other and thus, upon refolding, reduces the intermolecular interactions which lead to aggregation. The feasibility of this process was first demonstrated with Ribonuclease A as a model protein. In the present work, we expanded the scope of this study to better understand both the general mechanisms of protein refolding in reversed micelles and the biotechnological applicability of the process. First, we investigated the interactions between the individual components of the reversed micellar system (the protein molecule, the denaturant guanidine hydrochloride (GuHCl), and the surfactant (AOT)) during the refolding process. We then extended our studies to a more hydrophobic protein, γ‐interferon, which aggregates upon refolding in aqueous solution. However, it was also found to aggregate in our reversed micelle process during the extraction step. Since γ‐interferon is a much more hydrophobic protein than RNase, we hypothesize that interactions between hydrophobic amino acids and the surfactant layer may interfere with refolding. This hypothesis was tested by studying the refolding of chemically modified RNase. The substitution of 55% of the surface lysine residues with hydrophobic caproyl groups caused a significant decrease in the refolding yield of RNase in the reversed micellar system without affecting aqueous solution renaturation. In addition, the extraction efficiency of the enzyme from reversed micelles back into aqueous solution was severely reduced and resulted in aggregation. These experiments indicate that unfolded hydrophobic Proteinsinteract with the Surfactant molecules, which limits their ability to refold in reversed micelles.

Journal

Biotechnology and BioengineeringWiley

Published: Apr 25, 1990

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