Trinitrobenzenesulfonate modification of the lysine residues in lactose repressor protein.
Modification of the lysine residues in the lactose repressor protein has been carried out with trinitrobenzenesulfonate. Reaction of lysine residues at positions 33, 37, 108, 290, and 327 was observed. Inducer binding was increased by modification with this reagent, while both nonspecific DNA binding and operator DNA binding were diminished, although to differing degrees. The loss in operator DNA binding capacity was complete with modification of approximately 2 equiv of lysine per monomer. The extent of reaction was affected by the presence of both sugar and DNA ligands; binding activities of the modified protein and reaction pattern of the lysines were perturbed by these ligands. The presence of operator or nonspecific DNA during the reaction protected against specific and nonspecific DNA binding activity loss. This protection presumably occurs by steric restriction of reagent access to lysine residues which are essential for both nonspecific and operator binding interactions. Lysines-33 and -108 were protected from modification in the presence of DNA. These experiments suggest that the charge on the lysine residues is important for protein interaction with DNA and that steric constraints for operator DNA interaction with the protein are more restrictive than for nonspecific DNA binding. In contrast, inducer (isopropyl beta-D-thiogalactoside) presence partially protected lysine-290 from modification while significantly enhancing reaction at lysine-327. Conformational alterations consequent to inducer binding are apparently reflected in these altered lysine reactivities.
D002851:Chromatography, High Pressure Liquid; D004247:DNA / Q000378:metabolism; D004926:Escherichia coli / Q000032:analysis; D007544:Isopropyl Thiogalactoside / Q000494:pharmacology; D007763:Lac Operon; D008239:Lysine; D009578:Nitrobenzenes / Q000494:pharmacology*; D010446:Peptide Fragments; D011487:Protein Conformation; D012097:Repressor Proteins / Q000378:metabolism*; D013329:Structure-Activity Relationship; D014157:Transcription Factors / Q000378:metabolism*; D014302:Trinitrobenzenesulfonic Acid / Q000494:pharmacology*
D009578:Nitrobenzenes; D010446:Peptide Fragments; D012097:Repressor Proteins; D014157:Transcription Factors; D007544:Isopropyl Thiogalactoside; D014302:Trinitrobenzenesulfonic Acid; D004247:DNA; D008239:Lysine