Part A. Studies Directed Toward the Total Synthesis of Chebulagic Acid. Part B. DNA Recognition by Metallointercalator-Peptide Conjugates

Citation

Hastings, Curtis Asa (1998) Part A. Studies Directed Toward the Total Synthesis of Chebulagic Acid. Part B. DNA Recognition by Metallointercalator-Peptide Conjugates. Dissertation (Ph.D.), California Institute of Technology. doi:10.7907/e4be-bd57. https://resolver.caltech.edu/CaltechTHESIS:08082025-165811908

Abstract

Part A. The development of asymmetric allene/enone and allene/enoate intramolecular [2+2]-photocycloadditions is described. Irradiation of optically active allenes (89-92% ee) appended to enones and enoates afforded alkylidenecyclobutane photoadducts with high levels of asymmetric induction (83 -100%) derived exclusively from the allene fragment. Substrates examined include allenyl alcohols appended to 1,3- cyclopentanedione, 1,3-cyclohexanedione, and 4-hydroxycoumarin. The absolute sense of induction in these reactions was determined by photocycloaddition of an allene containing an internal stereochemical label. The exa-methylenecyclobutanes obtained upon irradiation of allene-coumarins were isolated as single olefin diastereomers. A model for the high levels of enantioinduction observed in these transformations is presented.

The asymmetric intramolecular allene/enoate [2+2]-photocycloaddition was applied to the synthesis of the topoisomerase I inhibitor chebulagic acid. The synthetic utility of this reaction was demonstrated by the preparation of an advanced intermediate containing all of the stereochemical information present in the chebulic acid fragment of chebulagic acid. In the course of these synthetic studies an unusual 8-alkynyl lactone photoproduct was obtained upon photocyloaddition of a substrate that was expected to afford a [7.6.6.4]tetracyclic system. A 1,5-hydrogen shift in a biradical intermediate was implicated in the formation of this product by isotopic labeling studies; the mechanistic implications of these results for enantioselectivity in photochemical reactions of optically active allenes tethered to enones and enoates are discussed.

Part B. The DNA recognition properties of peptide conjugates of phenenthrenequinone diimine complexes of rhodium(III) have been studied. The structural and thermodynamic basis for the 5'-CCA-3'-selectivity of the metallointercalator-peptide conjugate [Rh(phi)₂(phen')]³⁺ -AANVAIAAWERAA-CONH₂ was investigated. A protocol for measuring dissociation constants of DNA cleaving ligands by cleavage titration is described. Using this protocol, the energetic contribution of the peptide to sequenceselective binding was assessed, and evidence for the origin of the enhanced sequenceselectivity observed at elevated temperature was obtained. Micromolar quantities of [ΔRh-(phi)₂(phen')]³⁺-AANVAIAAWERAA-CONH₂ were synthesized to examine the structure of the metallointercalator-peptide conjugate and the metallointercalator-peptide conjugate•DNA complex by NMR. NMR results for the metallointercalator-peptide conjugate in the absence of DNA are reported.

A family of peptide conjugates of [Rh(phi)₂(phen')]³⁺(phi = 9, 10- phenanthrenequinone diimine, phen' = 5-(amidoglutaryl)-1,10-phenanthroline) was also synthesized. The peptide sequences were obtained by single amino acid modification of the 5'-CCA-3'-selective metallointercalator-peptide conjugate [Rh(phi)₂(phen')]³⁺- AANVAIAAWERAA-CONH₂ to explore the correlation between the amino acid sequence of the peptide and the nucleotide sequence of the DNA target. Changing the position of the glutamate at position 10 in the sequence of the appended peptide resulted in the identification of a 5'-ACA-3'-selective metallointercalator-peptide conjugate, [Rh(phi)₂(phen')³⁺-AANVAEAAWARAA-CONH₂. Locating the glutamate on one face of a putative α-helix was found to be essential for sequence specificity; peptide conjugates with the glutamate at positions 7, 8, 12, and 13 did not afford sequence-selective DNA recognition. Further amino acid substitutions were made at positions 6 and 10. Mutating the glutamate at position 6 to arginine caused complex changes in the recognition characteristics of the resulting conjugate. To probe the interactions that give rise to sequence specificity, we have measured thermodynamic dissociation constants for these sequence-selective metallointercalator-peptide conjugates and [Rh(phi)₂(phen')]³⁺.

The observed sequence preferences are consistent with the model of Sardesai et al. for the sequence selectivity of Rh(phi)₂(phen')]³⁺-AANVAIAAWERAA-CONH₂. This model states that sequence-specific DNA recognition requires the peptide to adopt an α-helical conformation, and that Glu¹⁰ makes a critical base-specific contact with the 5'- terminal cytosine of the recognition sequence. Using the additional sequence-selectivity data, this model is refined. This refined model suggests that recognition of the central C•G base pair of the 5'-CCA-3' recognition sequence of [Rh]-E10 is accomplished by Ile⁶ through shape-selection, and that recognition of the 5' -terminal A•T base pair of the 5' - ACA-3' recognition sequence of [Rh]-E6 is accomplished by van der Waals contacts between alanine and thymine methyl groups. The implications of these results for the de nova design of sequence-selective DNA binding peptides are discussed.

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