THE BACTERIAL PERIPLASMIC HISTIDINE-BINDING PROTEIN - STRUCTURE FUNCTION ANALYSIS OF THE LIGAND-BINDING SITE AND COMPARISON WITH RELATED PROTEINS/

Bacterial periplasmic binding proteins are initial receptors in the pr ocess of active transport across cell membranes and/or chemotaxis. Amo ng them, the histidine-binding protein (HisJ) has been extensively stu died from the biochemical, physiological, and genetic points of view. The three dimensional crystal structure of the histidine binding prote in complexed with histidine has been determined at 2.5-Angstrom resolu tion by the molecular replacement method using as a probe structure th e previously solved lysine liganded structure of the lysine-, arginine -, ornithine binding protein (LAG), which shares 70% sequence identity with HisJ. The structure is bi-lobate; the two lobes, one bigger than the other, are connected by two short strands and are in contact with each other (closed) enclosing the histidine. Charged, polar, and non- polar side chains, as well as the peptide backbone, are involved in ti ght binding of the histidine. The bound histidine is involved in eight direct hydrogen bonds, six with the bigger lobe and two with the smal ler lobe, in one potential water-mediated hydrogen bond with the bigge r lobe, as well as in ionic interactions. The HisJ residues surroundin g the ligand are the same as the LAO residues interacting with lysine, except for residue 52 which is leucine in HisJ and phenylalanine in L AG. The Leu-52 in HisJ makes a hydrophobic interaction with the imidaz ole ring of histidine. Of seven mutations affecting the ligand-binding site, five are located in the ligand-binding site, one in a con stran d, and one at the domains interface. Based on comparisons among relate d binding proteins, the specific interactions between the ligands and the respective binding protein residues are predicted for the glutamin e-binding protein and the opines-binding protein