Lactose repressor protein modified with dansyl chloride: activity effects and fluorescence properties.
Chemical modification using 5-(dimethylamino)naphthalene-1-sulfonyl chloride (dansyl chloride) has been used to explore the importance of lysine residues involved in the binding activities of the lactose repressor and to introduce a fluorescent probe into the protein. Dansyl chloride modification of lac repressor resulted in loss of operator DNA binding at low molar ratios of reagent/monomer. Loss of nonspecific DNA binding was observed only at higher molar ratios, while isopropyl beta-D-thiogalactoside binding was not affected at any of the reagent levels studied. Lysine residues were the only modified amino acids detected. Protection of lysines-33 and -37 from modification by the presence of nonspecific DNA correlated with maintenance of operator DNA binding activity, and reaction of lysine-37 paralleled operator binding activity loss. Energy transfer between dansyl incorporated in the core region of the repressor protein and tryptophan-201 was observed, with an approximate distance of 23 A calculated between these two moieties.
D002250:Carbon Radioisotopes; D004247:DNA / Q000378:metabolism; D003619:Dansyl Compounds / Q000494:pharmacology*; D004926:Escherichia coli / Q000378:metabolism*; D007544:Isopropyl Thiogalactoside / Q000494:pharmacology; D007700:Kinetics; D011485:Protein Binding; D012097:Repressor Proteins / Q000378:metabolism*; D013050:Spectrometry, Fluorescence; D014157:Transcription Factors / Q000378:metabolism*
D002250:Carbon Radioisotopes; D003619:Dansyl Compounds; D012097:Repressor Proteins; D014157:Transcription Factors; D007544:Isopropyl Thiogalactoside; D004247:DNA; C029038:dansyl chloride