Interdomain salt bridges modulate ligand-induced domain motion of the sulfate receptor protein for active transport.
The refined crystal structure of the liganded form of the Salmonella typhimurium sulfate-binding protein, a periplasmic receptor of active transport, is made up of two globular domains bisected by a deep cleft wherein the dehydrated sulfate is completely engulfed and bound by hydrogen bonds and van der Waals' forces. Two salt bridges (between Glu15 and Arg174 and between Asp68 and Arg134) span the cleft opening. To elucidate the role of the inter-domain salt bridges in the ligand-induced domain motion, the acidic residues were changed (singly and together) to their corresponding amide side-chains by site-directed mutagenesis of the recombinant Escherichia coli sulfate-binding protein. Rapid kinetics and equilibrium measurements of sulfate binding to the purified mutant proteins demonstrate that these salt bridges stabilize the closed liganded form of the receptor and modulate the rate of cleft opening. Our results have new implications in understanding the dynamics of many other multidomain proteins that undergo similar large-scale domain motions.
Jacobson
B L
BL
Department of Biochemistry, Baylor College of Medicine, Houston, TX 77030.
He
J J
JJ
Lemon
D D
DD
Quiocho
F A
FA
D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics; D001693:Biological Transport, Active; D002352:Carrier Proteins / Q000737:chemistry* / Q000235:genetics; D004252:DNA Mutational Analysis; D029968:Escherichia coli Proteins; D008024:Ligands; D046911:Macromolecular Substances; D033902:Periplasmic Binding Proteins; D011487:Protein Conformation; D011956:Receptors, Cell Surface / Q000737:chemistry / Q000235:genetics; D011994:Recombinant Proteins / Q000737:chemistry; D012486:Salmonella typhimurium / Q000502:physiology*; D012492:Salts / Q000737:chemistry; D013329:Structure-Activity Relationship
D001426:Bacterial Proteins; D002352:Carrier Proteins; D029968:Escherichia coli Proteins; D008024:Ligands; D046911:Macromolecular Substances; D033902:Periplasmic Binding Proteins; D011956:Receptors, Cell Surface; D011994:Recombinant Proteins; D012492:Salts; C489040:sbp protein, E coli; C044773:sulfate-binding protein, bacteria