Identification of a novel determinant for basic domain-leucine zipper DNA binding activity in the acute-phase inducible nuclear factor-interleukin-6 transcription factor

Allan R. Brasier, Amalendra Kumar

Research output: Contribution to journalArticle

17 Citations (Scopus)

Abstract

Nuclear factor-interleukin-6 (NF-IL6), a member of the CCAAT box/enhancer- binding protein (C/EBP) family, contains a basic domain-leucine zipper (bZIP) DNA binding motif. Controlled protease digestion was used to probe free and DNA-complexed NF-IL6 protein. Digestion with trypsin in the absence of DNA produced the leucine zipper domain (containing residues 303-345). In contrast, digestion of NF-IL6·DNA complexes produced a stable domain, spanning residues 266-345, termed the tryptic core domain (TCD). The NH2- terminal boundary of the TCD is longer than tryptic peptides reported from C/EBPα·DNA complexes. Digestion of NF-IL6 with endoprotease Asp-N produced a domain smaller than the TCD (NF-IL6 bZIP domains (NFBD) (272-345)), a domain identified either in the absence or the presence of DNA. Both recombinant peptides bind acute-phase response element DNA in a sequence- specific fashion. The equilibrium disassociation constant (K(d)) for the TCD was 36 ± 8 nM, whereas the K(d) for NFBD (272-345) was 283 ± 160 nM. Moreover, in comparison with the TCD, NFBD (272-345) formed unstable DNA complexes with a 15-fold faster off-rate. We conclude that the amino acids represented between 266 and 272 termed the complex stabilizing subdomain, influences DNA complex formation independent of DNA binding specificity, and may be one mechanism for heterogeneity of DNA interaction by C/EBP family members.

Original languageEnglish (US)
Pages (from-to)10341-10351
Number of pages11
JournalJournal of Biological Chemistry
Volume269
Issue number14
StatePublished - Apr 8 1994

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CCAAT-Enhancer-Binding Protein-beta
Leucine Zippers
Transcription Factors
DNA
Digestion
CCAAT-Enhancer-Binding Proteins
Carrier Proteins
Acute-Phase Reaction
Nucleotide Motifs
DNA Probes
Response Elements
Trypsin
Equilibrium constants
Peptide Hydrolases
Amino Acids
Peptides

ASJC Scopus subject areas

  • Biochemistry

Cite this

Identification of a novel determinant for basic domain-leucine zipper DNA binding activity in the acute-phase inducible nuclear factor-interleukin-6 transcription factor. / Brasier, Allan R.; Kumar, Amalendra.

In: Journal of Biological Chemistry, Vol. 269, No. 14, 08.04.1994, p. 10341-10351.

Research output: Contribution to journalArticle

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