The solution structure and dynamics of human neutrophil gelatinase-associated lipocalin

Human neutrophil gelatinase-associated lipocalin (HNGAL) is a member of the lipocalin family of extracellular proteins that function as transporters o f small, hydrophobic molecules. HNGAL, a component of human blood granulocy tes, binds bacterially derived formyl peptides that act as chemotactic agen ts and induce leukocyte granule discharge. HNGAL also forms a complex with the proenzyme form of matrix metalloproteinase-9 (pro-MMP-9, or progelatina se B) via an intermolecular disulphide bridge. This association allows the subsequent formation of ternary and quaternary metalloproteinase/inhibitor complexes that vary greatly in their metalloproteinase activities. The stru cture and dynamics of apo-HNGAL have been determined by NMR spectroscopy. S imulated annealing calculations yielded a set of 20 convergent structures w ith an average backbone RMSD from mean coordinate positions of 0.79(+/-0.13 ) Angstrom over secondary structure elements. The overall rotational correl ation time (13.3 ns) derived from N-15 relaxation data is consistent with a monomeric protein of the size of HNGAL (179 residues) under the experiment al conditions (1.4 mM protein, pH 6.0, 24.5 degrees C). The structure featu res an eight stranded antiparallel beta-barrel, typical of the lipocalin fa mily. One end of the barrel is open, providing access to the binding site w ithin the barrel cavity, while the other is closed by a short 3(10)-helix. The free cysteine residue required for association with pro-MMP-9 lies in a n inter-strand loop at the closed end of the barrel. The structure provides a detailed model of the ligand-binding site and has led to the proposal of a site for pro-MMP-9 association. Dynamic data correlate well with structu ral features, which has allowed us to investigate a mechanism by which a ce ll-surface receptor might distinguish between apo and holo-HNGAL through co nformational changes at the open end of the barrel. (C) 1999 Academic Press .