The long-term goal of this program is to determine the solution structure of site-specific and stereospecific adducts of styrene oxide in oligodeoxynucleotides, and to correlate this information with the results of adduct-directed mutagenesis and replication studies. the work will be done in the context of oligodeoxynucleotide duplexes which contain codons 11,12 and 13, 60, 61 and 62 of the human n-ras protooncogene. These are designated as the ras12 and ras61 sequences, respectively. Adduct-induced point mutations in codons 12 and 61 cause direct activation of this protooncogene. In the first phase of the work, already underway, """"""""primary"""""""" adduct structures will be examined. These are site-specific and stereospecific R/S-a-styrene oxide adducts which are base paired to the proper Watson-Crick complement. These structures provide information about styrene oxide-induced structural perturbations in duplex DNA. In the second phase of this work, """"""""secondary"""""""" adduct structures will be examined. These will be a series of postulated mutagenesis intermediates, designed to test hypotheses related to how specific R/S-a-styrene oxide adducts (a) are successfully bypassed in certain sequence and stereochemistry contexts but not in others, and (b) might induce activating point mutations in the ras12 and ras61 sequences. Studies underway focus on adduct of styrene oxide at the exocyclic amino groups of adenine and guanine: these will be continued. Structural studies of styrene oxide adducts at N7 of guanine will be initiated. High field NMR spectroscopy will be the primary technique used to obtain structural information. Structural refinement will be accomplished using molecular dynamics/simulated annealing algorithms which are restrained by combination of nuclear Overhauser (NOE)and scalar coupling data obtained from NMR.
Showing the most recent 10 out of 15 publications
