Experimental and Theoretical Studies of Charge-Charge Interactions in Proteins

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Understanding the mechanisms of stabilization of protein structure represents one of the majoraspects of the protein folding problem. The contributions of various short-range interactions,such as hydrogen bonding, the hydrophobic effect, and configurational entropy, to proteinstability have been well documented. However, it is becoming more and more evident that thelong-range electrostatic interactions between charged protein groups are also importantcontributors to protein structure and stability. The proposed work has an overall goal to explorethe relation between the charge-charge interactions on the protein surface and the proteinstability by engineering a thermostable protein by redesigning the charge-charge interactions on theprotein surface, and analyzing the optimized charge-charge interactions. The theoretical part of the work is based on the calculation of the charge-chargeinteractions in proteins using Tanford-Kirkwood approach with Bashford-Karplus approximation(TK-BK procedure). The experimental measurement of the stabilities of the protein variantsdesigned using TK-BK predictions will be done using differential scanning calorimetry, or usingchemical (urea) induced denaturation monitored by the changes in the circular dichroism orfluorescence intensity properties. The fundamental importance of this research is thatusing a combination of theoretical calculations and experimental measurements the relationshipbetween charge-charge interactions and the protein stability will be explored. From the practicalpoint this research will provide the foundation for the design of proteins stable at extremetemperatures. This will benefit the industrial use of proteins where thermostability of enzymes isin many cases the largest obstacle.