OPEN CONFORMATION OF A SUBSTRATE-BINDING CLEFT - F-19 NMR-STUDIES OF CLEFT ANGLE IN THE D-GALACTOSE CHEMOSENSORY RECEPTOR

The Escherichia coli D-galactose and D-glucose receptor is a two-domain structure with a sugar-binding site at the interface between domains. The structure of the closed cleft containing bound D-glucose has been determined crystallographically, but the open cleft remains to be characterized. The present study illustrates a generalizable approach that is used to detect and analyze both the open- and closed-cleft conformations in solution. A F-19 nucleus located inside the cleft is monitored by F-19 NMR. When the cleft is occupied by D-glucose, the F-19 nucleus is found to be inaccessible to the aqueous paramagnetic probe Gd.EDTA, verifying that the occupied cleft is closed in solution and inaccessible to bulk solvent. When the cleft is empty, the F-19 nucleus becomes accessible to the paramagnet such that the distance of closest approach is r less-than-or-equal-to 10 angstrom, indicating that the empty cleft opens at least transiently by an angle-theta greater-than-or-equal-to 18 +/- 3-degrees.