title: Multidimensional 1H and 15N NMR investigation of glutamine-binding protein of Escherichia coli.
abstract: Specific and uniform 15N labelings along with site-directed mutagenesis of glutamine-binding protein have been utilized to obtain assignments of the His156, Trp32 and Trp220 residues. These assignments have been made not only to further study the importance of these 3 amino acid residues in protein-ligand and protein-protein interactions associated with the active transport of L-glutamine across the cytoplasmic membrane of Escherichia coli, but also to serve as the starting points in the sequence-specific backbone assignment. The assignment of H epsilon 2 of His156 refines the earlier model where this particular proton forms an intermolecular hydrogen bond to the delta-carbonyl of L-glutamine, while assignments of both Trp32 and Trp220 show the variation in local structures which ensure the specificity in ligand binding and protein-protein interaction. Using 3D NOESY-HMQC NMR, amide connectivities can be traced along 8-9 amino acid residues at a time. This paper illustrates the usefulness of combining 15N isotopic labeling and multinuclear, multidimensional NMR techniques for a structural investigation of a protein with a molecular weight of 25,000.
authors.affiliation: Department of Biological Sciences, Carnegie Mellon University, Pittsburgh, PA 15213.
authors.forename: N
authors.initials: N
authors.lastname: Tjandra
authors.forename: V
authors.initials: V
authors.lastname: Simplaceanu
authors.forename: P F
authors.initials: PF
authors.lastname: Cottam
authors.forename: C
authors.initials: C
authors.lastname: Ho
mesh_terms: D000595:Amino Acid Sequence; D002352:Carrier Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D005973:Glutamine / Q000378:metabolism; D006859:Hydrogen; D009682:Magnetic Resonance Spectroscopy / Q000379:methods; D016297:Mutagenesis, Site-Directed; D009587:Nitrogen Isotopes; D011487:Protein Conformation
chemical_list: D002352:Carrier Proteins; D009587:Nitrogen Isotopes; C037853:glutamine transport proteins; D005973:Glutamine; D006859:Hydrogen