Conformational transitions of the lac repressor from Escherichia coli

Felicia Y.H. Wu*, Pradip Bandyopadhyay, Cheng Wen Wu

*Corresponding author for this work

Research output: Contribution to journalArticle

22 Scopus citations

Abstract

Temperature-jump studies of the lac repressor were performed monitoring the intrinsic tryptophan fluorescence of the protein. A single relaxation process with a time constant of 1·36×104 s-1 was observed for lac repressor solutions at pH 7·5 in the absence of inducer. The relaxation time is independent of the concentration of lac repressor indicating a conformational transition between two states of the repressor (R*g{cyrillic}R). Although the binding of an inducer, isopropyl-β-d-thiogalactoside, is too slow (as shown by stopped-flow studies) to influence the rate of the rapid interconversion between these two states of the lac repressor, the relaxation amplitude decreases with increasing IPTG concentration. The rate of this conformational transition is altered by protonation of the repressor. Temperature-jump data obtained for lac repressor at various pH values can be analyzed in terms of a concerted mechanism in which a proton binds preferentially to one of the two states of lac repressor (R). We propose that IPTG also binds selectively to the R state. This is consistent with our observation that addition of 10-3 m-IPTG shifts the pK of a specific group of amino acid residues in the lac repressor. Furthermore, the mechanism provides satisfactory explanations for previous reports that the bimolecular rate-constant for the IPTG-lac repressor interaction is extremely low and that the inducer binding may exhibit various degrees of co-operativity at different pH values. Thus the allosteric transition of lac repressor effected by proton and inducer may play a significant role in the regulation of lac transcription.†Abbriviation used: IPTG, isopropyl-β-D-thiogalactoside.

Original languageEnglish
Pages (from-to)459-472
Number of pages14
JournalJournal of Molecular Biology
Volume100
Issue number4
DOIs
StatePublished - 5 Feb 1976

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