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| The EMBO Journal
(1999) 18, 6271–6281, doi:10.1093/emboj/18.22.6271 |
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| N-terminal domain swapping and metal ion binding in nitric oxide synthase dimerization |
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| Brian R. Crane1, 2, Robin J. Rosenfeld1, Andrew S. Arvai1, Dipak K. Ghosh3, 4, Sanjay Ghosh3, John A. Tainer1, Dennis J. Stuehr3 and Elizabeth D. Getzoff1 |
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1 Department of Molecular Biology and The Skaggs Institute for Chemical Biology, The Scripps Research Institute, La Jolla, CA 92037, USA 2 Present address: The Beckman Institute, The California Institute of Technology, Pasadena, CA 91125, USA 3 Department of Immunology, The Cleveland Clinic, Cleveland, OH 44106, USA 4 Present address: Department of Medicine, Duke University and VA Medical Center, Durham, NC 27705, USA To whom correspondence should be addressed Brian R. Crane, crane@its.caltech.edu Elizabeth D. Getzoff, edg@scripps.edu Received 18 August 1999; Revised 30 September 1999; Accepted 30 September 1999. |
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| Abstract |
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Nitric oxide synthase oxygenase domains (NOSox) must bind tetrahydrobiopterin and dimerize to be active. New crystallographic structures of inducible NOSox reveal that conformational changes in a switch region (residues 103–111) preceding a pterin-binding segment exchange N-terminal  -hairpin hooks between subunits of the dimer. N-terminal hooks interact primarily with their own subunits in the 'unswapped' structure, and two switch region cysteines (104 and 109) from each subunit ligate a single zinc ion at the dimer interface. N-terminal hooks rearrange from intra- to intersubunit interactions in the 'swapped structure', and Cys109 forms a self-symmetric disulfide bond across the dimer interface. Subunit association and activity are adversely affected by mutations in the N-terminal hook that disrupt interactions across the dimer interface only in the swapped structure. Residue conservation and electrostatic potential at the NOSox molecular surface suggest likely interfaces outside the switch region for electron transfer from the NOS reductase domain. The correlation between three-dimensional domain swapping of the N-terminal hook and metal ion release with disulfide formation may impact inducible nitric oxide synthase (i)NOS stability and regulation in vivo. |
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| Keywords: disulfide, domain swapping, -hairpin, X-ray structure, zinc |
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