HIGH-RESOLUTION PROTON NUCLEAR MAGNETIC-RESONANCE STUDIES OF HISTIDINE-BINDING PROTEINS J OF SALMONELLA-TYPHIMURIUM - INVESTIGATION OF SUBSTRATE AND MEMBRANE INTERACTION SITES

The binding of l-histidine to the periplasmic histidine-binding proteins J of Salmonella typhimurium strains TA1859, TA301, and TA300 has been studied by high-resolution 1H nuclear magnetic resonance at 250 MHz. The dependence on substrate concentration of the chemical shifts of various resonances of each J protein was used to estimate a range of values for the off-rate constant k_1 for the binding of l-histidine to J protein. It was found that J proteins from two of the strains, TA1859 and TA300, have very similar values, while that from TA301 exhibits a much larger This indicates that the mutation in TA300 has not affected the l-histidine binding, but that the TA301 mutation has altered the conformation of the substrate binding site so that the substrate has a shorter bound lifetime, tb=1 jk_1. We have also observed differences between the spectra of the J protein from strain TA1859 (i. e., normal J protein) and the mutant J protein from TA300 or TA301. This shows that there are conformational differences between the native and mutant proteins, which have affected the environment of certain amino acid residues. From these observations we propose certain resonances as possibly arising from amino acid residues at particular regions of the J protein such as the substrate binding site or that part of the J protein molecule at which it interacts with the membrane-bound l-histidine permease or carrier. © 1979, American Chemical Society. All rights reserved.