The Human Genome Project is expected to uncover the genetic code for numerous proteins with an unknown structure or function. Therefore it has become increasingly more important to rapidly generate an active correctly folded protein form a given primary sequence. The use of recombinant expression systems in prokaryotic hosts has greatly simplified the large-scale production of eukaryotic proteins. A significant drawback to this otherwise powerful technique is that the protein of interest is often expressed in insoluble inclusion bodies. Inclusion bodies act to segregate the eukaryotic proteins into a misfolded or unfolded inactive state. Several methods have been developed to either minimize the formation of misfolded protein in vivo or to renature the protein in vitro into a correctly folded and active conformation. Although these strategies work to varying degrees on a case by case basis, there is no general method available for refolding denatured proteins. Our approach to this problem is the development of a series of alkylammonium salts and fluorous surfactants that not only prevent protein aggregation in a manner similar to common detergents, but also can be easily displaced from proteins thus enabling them to refold into their active conformations.
Our work is presently focused on two approaches to protein renaturation: 1) Development of a series of alkylammonium salts that not only prevent protein aggregation but also can be easily displaced from proteins thus enabling them to refold into their active conformation and, 2) Development of a series of fluorous surfactants that prevent aggregation of protein, but do not disrupt structure. The work within this second approach has two main goals: A) The development of a series of renaturation additives with varying degrees of hydrophobic and fluorophilic character, and B) The application of fluorous biphasic separation in the displacement of fluorous surfactants and additives from proteins.