Nuclear magnetic resonance and fluorescence studies of substrate-induced conformational changes of histidine-binding protein J of Salmonella typhimurium.

Abstract

The histidine-binding protein J of Salmonella typhimurium binds L-histidine as a first step in the high-affinity active transport of this amino acid across the cytoplasmic membrane. High-resolution nuclear magnetic resonance spectroscopy has been used to monitor the conformation of histidine-binding protein J in the presence and absence of substrate. Evidence is presented to show that this binding protein undergoes a conformational change involving a substantial number of amino-acid residues (including tryptophans) in the presence of L-histidine and that this change is specific for L-histidine. In order to monitor the involvement of tryptophan residues in the substrate-induced conformational change, 5-fluorotryptophan has been incorporated biosynthetically into the histidine-binding protein J using a tryptophan autotroph of Salmonella typhimurium. There are no significant differences in the conformation and binding activity between the 5-fluorotryptophan-labeled and the normal histidine-binding protein J. Proton and fluorine-19 nuclear magnetic resonance studies of the 5-fluorotryptophan-labeled binding protein show that at least one (and possibly two) of the tryptophan residues undergo(es) a change toward a more hydrophobic environment in the presence of L-histidine. These observations are supported by fluorescence data and by differences in the reactivity of the tryptophan residues of this protein toward N-bromosuccinimide in the presence and absence of substrate. The present results are consistent with models for the action of periplasmic-binding proteins in shock-sensitive transport systems of gram-negative bacteria which require a substrate-induced conformational change prior to the energy-dependent translocation of substrates.

• Authors: D E Robertson, P A Kroon, C Ho
• Journal: Biochemistry (Biochemistry) Vol. 16 Issue 7 Pg. 1443-51 (Apr 05 1977) ISSN: 0006-2960 [Print] United States
• PMID: 321019 (Publication Type: Journal Article, Research Support, U.S. Gov't, Non-P.H.S., Research Support, U.S. Gov't, P.H.S.)

Chemical References

• Carrier Proteins
• Ligands
• Histidine
• Tryptophan
• Bromosuccinimide

Topics

• Binding Sites
• Bromosuccinimide
• Carrier Proteins (isolation & purification, metabolism)
• Histidine (metabolism)
• Ligands
• Magnetic Resonance Spectroscopy
• Protein Binding
• Protein Conformation
• Salmonella typhimurium (metabolism)
• Spectrometry, Fluorescence
• Tryptophan (analysis)