Biomaterial Database

Ligand-induced conformational change in the ferric enterobactin receptor FepA as studied by site-directed spin labeling and time-domain ESR. 1998

C S Klug, and S S Eaton, and G R Eaton, and J B Feix

Biophysics Research Institute, Medical College of Wisconsin, Milwaukee 53226, USA.

A mutant of the ferric enterobactin receptor, FepA, containing a valine to cysteine (V338C) substitution was made and the purified protein selectively modified with a sulfhydryl-specific nitroxide spin label. In reconstituted liposomes, interaction of the attached spin label with a combination of water-soluble and lipid-soluble relaxation agents indicated that the V338C site was located in the polar headgroup region of the membrane, approximately 1.5-4.5 A above the phosphate groups of the lipids. Binding of the ligand, ferric enterobactin (FeEnt), to the purified spin-labeled protein produced a significant decrease in both the rotational freedom and accessibility of the nitroxide, indicating the formation of new structural contacts between the spin label and either the protein or the bound ligand. Electron spin-echo (ESE) measurements of the nitroxide phase-memory relaxation rate in the presence and absence of bound ligand showed substantial dipolar coupling between the Fe3+ of FeEnt and the spin label and provided an iron-nitroxide distance estimate in the range of 20-30 A. We conclude that the ligand-induced changes in spin label motion and accessibility are due to new tertiary contacts with the protein and not to direct contact with the ligand. These studies suggest that V338C may occupy a hinge region connecting the ligand binding surface loop to the beta-barrel and provide the strongest evidence to date of an in vitro ligand-induced conformational change in FepA.

UI	MeSH Term	Description	Entries
D008024	Ligands	A molecule that binds to another molecule, used especially to refer to a small molecule that binds specifically to a larger molecule, e.g., an antigen binding to an antibody, a hormone or neurotransmitter binding to a receptor, or a substrate or allosteric effector binding to an enzyme. Ligands are also molecules that donate or accept a pair of electrons to form a coordinate covalent bond with the central metal atom of a coordination complex. (From Dorland, 27th ed)	Ligand
D008051	Lipid Bilayers	Layers of lipid molecules which are two molecules thick. Bilayer systems are frequently studied as models of biological membranes.	Bilayers, Lipid,Bilayer, Lipid,Lipid Bilayer
D008081	Liposomes	Artificial, single or multilaminar vesicles (made from lecithins or other lipids) that are used for the delivery of a variety of biological molecules or molecular complexes to cells, for example, drug delivery and gene transfer. They are also used to study membranes and membrane proteins.	Niosomes,Transferosomes,Ultradeformable Liposomes,Liposomes, Ultra-deformable,Liposome,Liposome, Ultra-deformable,Liposome, Ultradeformable,Liposomes, Ultra deformable,Liposomes, Ultradeformable,Niosome,Transferosome,Ultra-deformable Liposome,Ultra-deformable Liposomes,Ultradeformable Liposome
D011487	Protein Conformation	The characteristic 3-dimensional shape of a protein, including the secondary, supersecondary (motifs), tertiary (domains) and quaternary structure of the peptide chain. PROTEIN STRUCTURE, QUATERNARY describes the conformation assumed by multimeric proteins (aggregates of more than one polypeptide chain).	Conformation, Protein,Conformations, Protein,Protein Conformations
D011956	Receptors, Cell Surface	Cell surface proteins that bind signalling molecules external to the cell with high affinity and convert this extracellular event into one or more intracellular signals that alter the behavior of the target cell (From Alberts, Molecular Biology of the Cell, 2nd ed, pp693-5). Cell surface receptors, unlike enzymes, do not chemically alter their ligands.	Cell Surface Receptor,Cell Surface Receptors,Hormone Receptors, Cell Surface,Receptors, Endogenous Substances,Cell Surface Hormone Receptors,Endogenous Substances Receptors,Receptor, Cell Surface,Surface Receptor, Cell
D002352	Carrier Proteins	Proteins that bind or transport specific substances in the blood, within the cell, or across cell membranes.	Binding Proteins,Carrier Protein,Transport Protein,Transport Proteins,Binding Protein,Protein, Carrier,Proteins, Carrier
D003545	Cysteine	A thiol-containing non-essential amino acid that is oxidized to form CYSTINE.	Cysteine Hydrochloride,Half-Cystine,L-Cysteine,Zinc Cysteinate,Half Cystine,L Cysteine
D004578	Electron Spin Resonance Spectroscopy	A technique applicable to the wide variety of substances which exhibit paramagnetism because of the magnetic moments of unpaired electrons. The spectra are useful for detection and identification, for determination of electron structure, for study of interactions between molecules, and for measurement of nuclear spins and moments. (From McGraw-Hill Encyclopedia of Science and Technology, 7th edition) Electron nuclear double resonance (ENDOR) spectroscopy is a variant of the technique which can give enhanced resolution. Electron spin resonance analysis can now be used in vivo, including imaging applications such as MAGNETIC RESONANCE IMAGING.	ENDOR,Electron Nuclear Double Resonance,Electron Paramagnetic Resonance,Paramagnetic Resonance,Electron Spin Resonance,Paramagnetic Resonance, Electron,Resonance, Electron Paramagnetic,Resonance, Electron Spin,Resonance, Paramagnetic
D004758	Enterobactin	An iron-binding cyclic trimer of 2,3-dihydroxy-N-benzoyl-L-serine. It is produced by E COLI and other enteric bacteria.	Enterochelin
D001425	Bacterial Outer Membrane Proteins	Proteins isolated from the outer membrane of Gram-negative bacteria.	OMP Proteins,Outer Membrane Proteins, Bacterial,Outer Membrane Lipoproteins, Bacterial