PHYSICOCHEMICAL STUDIES OF BIOCHEMICAL REACTIONS

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The broad aim in this project is the study of biological and biochemical processes using physicochemical techniques. As in the past, our efforts will be centered on the determination, primarily by calorimetric methods and the interpretation of the thermodynamics of such processes. A major fraction of our work will involve study of thermally-induced processes in proteins, nucleic acids and lipids by means of high sensitivity differential scanning calorimetry (DSC). The power and applicability of this technique is at present rapidly expanding. For example, DSC affords the best means for checking the thermal stability of proteins engineered to perform some specific pharmaceutical function, and for acquiring thermodynamic data which may suggest specific ways to improve the thermal stability. Our DSC work on proteins will remain centered on studying the effects of mutations using materials supplied to use by a wide range of collaborators. Included will be, among others, staphylococcal nuclease, streptomyces subtilisin inhibitor (SSI), thioredoxin and cytochrome b562. DSC work on nucleotides will include study of the unfolding of synthetic triple stranded DNA, and of various nucleotide-protein complexes also studies by ITC. DSC gives an excellent means for studying the phase transitions of lipids. Work on pure synthetic lipids of unusual structure will continue. Studies of the effects of added materials (Ca++, Mg++, AlCl3, nitrophenyl esters) on the thermotropic behavior of lipid bilayers will also be continued. We are making constantly increasing use of isothermal situation calorimetry (ITC), in studying the binding of ligands to proteins and nucleic acids, the interactions of proteins with nucleic acids, for example, the trp repressor-trp operator interaction, and other biochemical reactions.