title: Chemical modification of lactose repressor protein using N-substituted maleimides.
abstract: Lactose repressor protein has been modified with N-ethylmaleimide, two N-maleimide spin labels, and an N-maleimide fluorophore. The reaction with repressor cysteine residues has been characterized. Approximately 2 of the 3 eq of cysteine/repressor monomer are reactive toward these reagents. Repressor cysteines are reactive toward these reagents in the order cysteine 140 greater than or equal to cysteine 107 greater than cysteine 281. The reaction is sulfhydryl-specific. Comparison of chemical modification data obtained in this laboratory using a variety of sulfhydryl-specific reagents has been used to assess chemical features of individual cysteine environments. Effects of the maleimide reagents on biological activity have been determined. Only the fluorophore N-(3-pyrene)maleimide has significant effect; this agent selectively perturbs repressor's ability to bind to operator DNA. This result suggests that regions of protein structure surrounding 1 or more of the cysteine residues possess determinants required for normal operator DNA binding.
mesh_terms: D003545:Cysteine / Q000032:analysis; D004926:Escherichia coli; D005033:Ethylmaleimide; D008301:Maleimides; D011485:Protein Binding; D011487:Protein Conformation; D012097:Repressor Proteins; D013329:Structure-Activity Relationship; D014157:Transcription Factors
chemical_list: D008301:Maleimides; D012097:Repressor Proteins; D014157:Transcription Factors; D003545:Cysteine; D005033:Ethylmaleimide