NMR-derived model of interconverting conformations of an ICAM-1 inhibitory cyclic nonapeptide.
We have produced by phage-display a disulfide-linked cyclic nonapeptide (inhibitory peptide-01, IP01), CLLRMRSIC, that binds to intracellular adhesion molecule-1 (ICAM-1) and blocks binding to its counter-structure, leukocyte functional antigen-1 (LFA-1). As a first step towards improving its pharmacologic properties, we have performed a structural and functional analysis of this peptide inhibitor to determine the features relevant to ICAM-1 binding. We report here the solution model of our initial product, IP01, as derived from two-dimensional nuclear magnetic resonance (NMR) restraints and molecular modeling. Distance and dihedral angle restraints, generated from nuclear Overhauser effect spectroscopy (NOESY) and one-dimensional-NMR experiments respectively, were used to generate an ensemble of structures using distance geometry and simulated annealing. Molecular dynamic simulations produced three interconverting conformational families consistent with the NMR-derived constraints. We describe these conformations and their mechanism of interconversion. Furthermore, we have measured the IC50 s of a series of inhibitors generated from IP01 through alanine substitution of each residue. These results show that the L2-L3-R4-M5-R6 segment is functionally active, conformationally flexible, and contains a beta-turn involving residues R4-S7, while the C1-C9-I8-S7 segment is less functionally-active but adopts a more defined solution conformation, consistent with a scaffolding function. This model will be useful for designing nonpeptide-based organic inhibitors with improved pharmacologic properties.
Sillerud
L O
LO
Department of Biochemistry and Molecular Biology, University of New Mexico Health Sciences Center, Albuquerque, NM 87131, USA.
Burks
E J
EJ
Wester
M J
MJ
Brown
D C
DC
Vijayan
S
S
Larson
R S
RS
D002352:Carrier Proteins / Q000037:antagonists & inhibitors / Q000737:chemistry*; D018799:Intercellular Adhesion Molecule-1 / Q000737:chemistry*; D016169:Lymphocyte Function-Associated Antigen-1 / Q000737:chemistry; D009682:Magnetic Resonance Spectroscopy; D008958:Models, Molecular; D008968:Molecular Conformation; D010456:Peptides, Cyclic / Q000037:antagonists & inhibitors / Q000737:chemistry*; D011487:Protein Conformation
D002352:Carrier Proteins; D016169:Lymphocyte Function-Associated Antigen-1; D010456:Peptides, Cyclic; C473032:cyclic nonapeptide IP25; D018799:Intercellular Adhesion Molecule-1