TY - JOUR
T1 - Heme distortion modulated by ligand-protein interactions in inducible nitric-oxide synthase
AU - Lis, David
AU - Stuehr, Dennis J.
AU - Yeh, Syun Ru
AU - Rousseau, Denis L.
PY - 2004/6/18
Y1 - 2004/6/18
N2 - The catalytic center of nitric-oxide synthase (NOS) consists of a thiolate-coordinated heme macrocycle, a tetrahydrobiopterin (H4B) cofactor, and an L-arginine (L-Arg)/N-hydroxyarginine substrate binding site. To determine how the interplay between the cofactor, the substrates, and the protein matrix housing the heme regulates the enzymatic activity of NOS, the CO-, NO-, and CN-bound adducts of the oxygenase domain of the inducible isoform of NOS (iNOSoxy) were examined with resonance Raman spectroscopy. The Raman data of the CO-bound ferrous protein demonstrated that the presence of L-Arg causes the Fe-C-O moiety to adopt a bent structure because of an H-bonding interaction whereas H4B binding exerts no effect. Similar behavior was found in the CN-bound ferric protein and in the nitric oxide (NO)-bound ferrous protein. In contrast, in the NO-bound ferric complexes, the addition of L-Arg alone does not affect the structural properties of the Fe-N-O moiety, but H4B binding forces it to adopt a bent structure, which is further enhanced by the subsequent addition of L-Arg. The differential interactions between the various heme ligands and the protein matrix in response to L-Arg and/or H4B binding is coupled to heme distortions, as reflected by the development of a variety of out-of-plane heme modes in the low frequency Raman spectra. The extent and symmetry of heme deformation modulated by ligand, substrate, and cofactor binding may provide important control over the catalytic and autoinhibitory properties of the enzyme.
AB - The catalytic center of nitric-oxide synthase (NOS) consists of a thiolate-coordinated heme macrocycle, a tetrahydrobiopterin (H4B) cofactor, and an L-arginine (L-Arg)/N-hydroxyarginine substrate binding site. To determine how the interplay between the cofactor, the substrates, and the protein matrix housing the heme regulates the enzymatic activity of NOS, the CO-, NO-, and CN-bound adducts of the oxygenase domain of the inducible isoform of NOS (iNOSoxy) were examined with resonance Raman spectroscopy. The Raman data of the CO-bound ferrous protein demonstrated that the presence of L-Arg causes the Fe-C-O moiety to adopt a bent structure because of an H-bonding interaction whereas H4B binding exerts no effect. Similar behavior was found in the CN-bound ferric protein and in the nitric oxide (NO)-bound ferrous protein. In contrast, in the NO-bound ferric complexes, the addition of L-Arg alone does not affect the structural properties of the Fe-N-O moiety, but H4B binding forces it to adopt a bent structure, which is further enhanced by the subsequent addition of L-Arg. The differential interactions between the various heme ligands and the protein matrix in response to L-Arg and/or H4B binding is coupled to heme distortions, as reflected by the development of a variety of out-of-plane heme modes in the low frequency Raman spectra. The extent and symmetry of heme deformation modulated by ligand, substrate, and cofactor binding may provide important control over the catalytic and autoinhibitory properties of the enzyme.
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U2 - 10.1074/jbc.M400968200
DO - 10.1074/jbc.M400968200
M3 - Article
C2 - 15066989
AN - SCOPUS:2942755861
SN - 0021-9258
VL - 279
SP - 26489
EP - 26499
JO - Journal of Biological Chemistry
JF - Journal of Biological Chemistry
IS - 25
ER -