The objective of this project is to extend the applications of optical spectroscopy, especially circular dichroism (CD), in the study of macromolecular structure and function. The general approach is to combine experimental and theoretical methods wherever practical, and to study both model systems and systems of direct biological interest. During the next five years three major areas will be studied: (1) the intrinsic CD of proteins, with studies of both backbone and sidechain contributions; (2) extrinsic CD contributions of chromophoric prosthetic groups and coenzymes; (3) the CD of nucleic acids. The current model for calculating polypeptide CD will be refined. The improved model will be applied to beta-turns and betasheets, and unordered peptides. Interactions among alpha helices and between alpha helices and beta-sheets in supersecondary structures will be considered. The extent to which end effects influence the magnitude of the alpha-helix CD spectrum will be assessed experimentally by studies of short peptides designed to have enhanced alpha-helical stability. A general program to calculate side-chain CD contributions in proteins will be developed, including the three aromatic side chains, histidine and disulfides. This program will be applied to a series of proteins which are known to have anomalous far ultraviolet CD. Calculations will also be performed for bovine pancreatic trypsin inhibitor, in which a series of site-directed mutations have replaced aromatic groups or disulfides. The effects of protein dynamics on sidechain and backbone CD will be investigated by combining CD calculations with MD simulations on small proteins. A cluster of four tyrosyl residues in tropomyosin will also be modeled by MD and the side-chain CD contributions will be calculated. The role of deviations from planarity in the heme of various heme proteins in determining the CD will be explored. The effects of site-directed mutations in the reaction center of a photosynthetic bacterium will be studied both experimentally and theoretically. Crystal field effects on the charge distribution and electronic spectra of the bases will be calculated. The calculations will then be extended to nucleosides, nucleotides and oligonucleotides. The methods developed for the crystals will be applied to calculations on oligonucleotides in solution, including both absorption and CD.
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