Two systems for studying the specific interaction of proteins with duplex DNA are under study by a combination of biochemical and genetic techniques. The first, known as the trpDll system, focuses upon a short stretch of DNA within the trp operon of E. coli which can gain or lose promoter activity by single base pair changes. Of twenty different alleles of trpDll which have been isolated, all affecting the same codon, fourteen exhibit full promoter activity while the remaining six are inactive. Our work is directed toward determining the location of trpDll mutations by sequencing cloned DNA using the Maxam-Gilbert method. In a complementary study we will employ an in vivo transcription system to ascertain which parameters of RNA polymerase-promoter interaction are affected by mutations within the trpDll locus. The second system under study is the trpR-Trp repressor gene-protein system. The structure of the trpR gene and its protein product are known, although the repressor has not yet been prepared in pure form. By suitable manipulation of plasmids harboring the trpR gene we will develop a large-scale production method for the protein product, then study by direct chemical means its interaction with trp operator.
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