Cited by 34Nitric Oxide and Endothelin EffectsHemoglobin structure and functionMetal-Catalyzed Oxygenation Mechanisms

Versatile Regulation of Neuronal Nitric Oxide Synthase by Specific Regions of Its C-Terminal Tail

Tiso, Mauro, Tejero, Jesús, Panda, Koustubh, Aulak, Kulwant S., Stuehr, Dennis J.·2007·Biochemistry

Abstract

The C-terminal tail (CT) of neuronal nitric oxide synthase (nNOS) is a regulatory element that suppresses nNOS activities in the absence of bound calmodulin (CaM). A crystal structure of the nNOS reductase domain (nNOSr) (Garcin, E. D., Bruns, C. M., Lloyd, S. J., Hosfield, D. J., Tiso, M., Gachhui, R., Stuehr, D. J., Tainer, J. A., and Getzoff, E. D. (2004) J. Biol. Chem. 279, 37918-37927) revealed how the first half of the CT interacts with nNOSr and thus provided a template for detailed studies. We generated truncation mutants in nNOS and nNOSr to test the importance of 3 different regions of the CT. Eliminating the terminal half of the CT (all residues from Ile1413 to Ser1429), which is invisible in the crystal structure, had almost no impact on NADP+ release, flavin reduction, flavin autoxidation, heme reduction, reductase activity, or NO synthesis activity, but did prevent an increase in FMN shielding that normally occurs in response to NADPH binding. Additional removal of the CT alpha-helix (residues 1401 to 1412) significantly increased the NADP+ release rate, flavin autoxidation, and NADPH oxidase activity, and caused hyper-deshielding of the FMN cofactor. These effects were associated with increased reductase activity and slightly diminished heme reduction and NO synthesis. Further removal of residues downstream from Gly1396 (a full CT truncation) amplified the aforementioned effects and in addition altered NADP+ interaction with FAD, relieved the kinetic suppression on flavin reduction, and further diminished heme reduction and NO synthesis. Our results reveal that the CT exerts both multifaceted and regiospecific effects on catalytic activities and related behaviors, and thus provide new insights into mechanisms that regulate nNOS catalysis.

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Crossref · 10.1021/bi701646k

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Authors
Tiso, M., Tejero, J., Panda, K., Aulak, K. S., & Stuehr, D. J.
Affiliations
Cleveland State University (US)
Corresponding author
  • Tiso, Mauro
  • Stuehr, Dennis J.
Venue
Biochemistry
Published
2007-11-20
Volume
46
Issue
50
Pages
14418-14428
ISSN
  • 0006-2960 (ISSN-L)
  • 1520-4995
OpenAlex
W1969901042
Cited by
34
References
64
Citations by year
2026: 2 · 2018: 5 · 2016: 3 · 2014: 4 · 2013: 4 · 2012: 4
FWCI
2.45
Type
journal article
Language
en
Open access
closed
Locations
  • Biochemistry (publishedVersion)
  • PubMed (publishedVersion)
Funders
  • National Heart, Lung, and Blood Institute (HL76491, P01HL076491)
  • National Institute of General Medical Sciences (GM51491, R01GM051491)
Topics
  • Health Sciences › Medicine › Physiology › Nitric Oxide and Endothelin Effects
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Cell Biology › Hemoglobin structure and function
  • Physical Sciences › Chemistry › Inorganic Chemistry › Metal-Catalyzed Oxygenation Mechanisms
Keywords
neuronal nitric oxide synthase, C-terminal tail, calmodulin, enzyme regulation, heme reduction, NADPH oxidase activity, truncation mutants, catalytic mechanism
MeSH terms
Amino Acid Sequence, Animals, Computer Simulation, Cytochrome Reductases, Flavin-Adenine Dinucleotide, Flavin Mononucleotide, Heme, Kinetics, Molecular Sequence Data, Mutation, NADP, Oxidation-Reduction, Protein Binding, Protein Structure, Secondary, Protein Structure, Tertiary, Rats, Nitric Oxide Synthase Type I

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