This application outlines research which focuses on the use of molecular micelles to replace sodium dodecyl sulfate (SDS) and other conventional micelles in poly- acrylamide gel electrophoresis. The overall hypothesis of this research stems from our belief that molecular micelles should provide better PAGE separation of proteins than SDS or other conventional micelles. This hypothesis is based upon previous separation strategies developed in our laboratory for separation of chiral molecules. In addition, we have preliminary data provided in this application which supports this hypothesis. Thus, the overall objective of the research described here is to examine better separation strategies for analyses of protein samples using molecular micellar-polyacrylamide gel electrophoresis (MoMi-PAGE). To achieve this objective, this application is divided into three specific aims. The first specific aim is to understand the parameters which contribute to PAGE separations using molecular micelles as alternative reagents to SDS and other conventional micelles which are frequently used in PAGE separations. This first specific aim builds upon expertise developed over the past several years in our laboratory. Under this specific aim, we will explore various strategies with molecular micelles of specifically designed architectures to optimize separations for different classes of proteins using PAGE separations. Our second specific aim is to understand the mechanisms of our separations at the molecular level since previous success with our separation research has been the result of treating our separation problems as a molecular recognition problem. To achieve this aim, we will use fluorescent probes and experimental design to understand these parameters which contribute to optimal separations. Our third specific aim is to apply the technology that we have developed to the analyses of proteins associated with the formation of atherosclerotic plaques and proteins associated with other biological systems of interest. This latter specific aim is mostly descriptive of the kinds of application that we will ultimately explore since it is unlikely that we would get to such applications during the proposed two-year time frame of this application. Finally, it is noted that our previous studies under NIH funding, as well as the individual expertise of the collaborators brought together for this research, enhances the chance for success of the overall goals outlined in this application. Protein separations are very important to understanding many diseases which affect human health. Therefore, it is important to develop better separation and identification procedures for analyses of proteins. The most widely used protein separation technique is called sodium dodecyl sulfate (SDS) polyacrylamide gel electrophoresis (PAGE) or (SDS-PAGE), which employs SDS for solubilization and separation of proteins on an electric grid. We propose to develop new reagents, call molecular micelles, to replace SDS and other surfactants in order to improve PAGE separation of proteins.
| Das, Susmita; Sylvain, Monica R; Fernand, Vivian E et al. (2011) Positive cooperative mechanistic binding of proteins at low concentrations: a comparison of poly (sodium N-undecanoyl sulfate) and sodium dodecyl sulfate. J Colloid Interface Sci 363:585-94 |
