DOI: 10.5176/2251-2489_BioTech47
Authors: Afra Alblooshi, Ibtesam Saeed and Syed Ashraf
Abstract: Green Fluorescent Protein (GFP) is an unusually stable auto-fluorescent protein that is widely used for various life sciences and biotechnology applications. (The importance of GFP can be judged from the fact that the 2008 Nobel prize in Chemistry was awarded for the discovery and applications of GFP). We have previously shown that the unusual stability attributed to GFP only hold true at pH values higher than 7. At pH values of 6.5 or lower, GFP dramatically loses its robust stability and becomes very susceptible to various denaturants such as heat or detergents. In the present study, we present additional data showing that during the denaturation of GFP, the protein may undergo through stable partially denatured states (molten globules). We believe that we have identified conditions that can allow us to generate these molten globules and study the unique fluorophore of GFP as well as its secondary and tertiary structures. We have used the “Red Edge Excitation Shift, REES” approach to further study the GFP fluorophore and gain valuable information about its dynamics and orientation inside the β-barrel structure of GFP. Studies like these will surely be of much interest to scientists studying GFP and will lead to yet more undiscovered applications of this very versatile and useful protein.
Keywords: Green Fluorescent Protein; GFP; thermal denaturation; SDS; Red Edge Excitation Shift; REES; moten globules; stable intermediates
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