Multiphasic denaturation of the λ repressor by urea and its implications for the repressor structure

• Published in: European journal of biochemistry Vol. 206; no. 1; pp. 15 - 21
• Main Authors: BANIK, Utpal, SAHA, Rina, MANDAL, Nitai C., BHATTACHARYYA, Bhabatarak, ROY, Siddhartha
• Format: Journal Article
• Language: English
• Published: Oxford, UK Blackwell Publishing Ltd 15-05-1992; Blackwell
• Subjects: Biological and medical sciences; DNA-Binding Proteins; Escherichia coli - metabolism; Fundamental and applied biological sciences. Psychology; Kinetics; Molecular and cellular biology; Molecular genetics; Peptide Fragments - isolation & purification; Protein Conformation; Protein Denaturation; Repressor Proteins - chemistry; Repressor Proteins - drug effects; Spectrometry, Fluorescence; Sulfhydryl Compounds - metabolism; Transcription Factors - chemistry; Transcription. Transcription factor. Splicing. Rna processing; Tryptophan; Urea - pharmacology; Viral Proteins; Viral Regulatory and Accessory Proteins; Urea; Transcription repressor; Chemical modification; Molecular structure; Structure function relationship
• ISSN: 0014-2956; 1432-1033
• DOI: 10.1111/j.1432-1033.1992.tb16896.x

Urea denaturation of the λ repressor has been studied by fluorescence and circular dichroic spectroscopies. Three phases of denaturation could be detected which we have assigned to part of the C‐terminal domain, N‐terminal domain and subunit dissociation coupled with further denaturation of the rest of the C‐terminal domain at increasing urea concentrations. Acrylamide quenching suggests that at least one of the three tryptophan residues of the λ repressor is in a different environment and its emission maximum is considerably blue‐shifted. The transition in low urea concentration (midpoint approximately 2 M) affects the environment of this tryptophan residue, which is located in the C‐terminal domain. Removal of the hinge and the N‐terminal domain shifts this transition towards even lower urea concentrations, indicating the presence of interaction between hinge on N‐terminal and C‐terminal domains in the intact repressor.