Chimeras of nitric-oxide synthase types I and III establish fundamental correlates between heme reduction, heme-NO complex formation, and catalytic activity

The Journal of Biological Chemistry
S AdakD J Stuehr

Abstract

Neuronal nitric-oxide synthase (nNOS or NOS I) and endothelial NOS (eNOS or NOS III) differ widely in their reductase and nitric oxide (NO) synthesis activities, electron transfer rates, and propensities to form a heme-NO complex during catalysis. We generated chimeras by swapping eNOS and nNOS oxygenase domains to understand the basis for these differences and to identify structural elements that determine their catalytic behaviors. Swapping oxygenase domains did not alter domain-specific catalytic functions (cytochrome c reduction or H(2)O(2)-supported N(omega)-hydroxy-l-arginine oxidation) but markedly affected steady-state NO synthesis and NADPH oxidation compared with native eNOS and nNOS. Stopped-flow analysis showed that reductase domains either maintained (nNOS) or slightly exceeded (eNOS) their native rates of heme reduction in each chimera. Heme reduction rates were found to correlate with the initial rates of NADPH oxidation and heme-NO complex formation, with the percentage of heme-NO complex attained during the steady state, and with NO synthesis activity. Oxygenase domain identity influenced these parameters to a lesser degree. We conclude: 1) Heme reduction rates in nNOS and eNOS are controlled primarily by their...Continue Reading

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Citations

Dec 4, 2012·International Journal of Molecular Sciences·Louise AigrainGilles Truan
Jan 5, 2002·Proceedings of the National Academy of Sciences of the United States of America·Subrata AdakDennis J Stuehr
Oct 3, 2002·Proceedings of the National Academy of Sciences of the United States of America·Subrata AdakDennis J Stuehr
May 23, 2007·Proceedings of the National Academy of Sciences of the United States of America·Mohammad Mahfuzul HaqueDennis J Stuehr
Aug 16, 2014·Proceedings of the National Academy of Sciences of the United States of America·Melody G CampbellMichael A Marletta
Mar 13, 2014·The Journal of Biological Chemistry·Anna Sobolewska-StawiarzNigel S Scrutton
Jan 23, 2015·The Biochemical Journal·Luciana HannibalDennis J Stuehr
Mar 4, 2015·Biochemical and Biophysical Research Communications·Rekha KarLinda J Roman
Mar 31, 2015·PloS One·Latika Nagpal, Koustubh Panda
Feb 8, 2018·The Journal of Biological Chemistry·Mohammad Mahfuzul HaqueDennis J Stuehr
Jun 6, 2002·The Journal of Biological Chemistry·Koustubh PandaDennis J Stuehr
May 11, 2004·The Journal of Biological Chemistry·Dennis J StuehrSubrata Adak
Sep 10, 2005·The Journal of Biological Chemistry·Mauro TisoDennis J Stuehr
Sep 27, 2006·The Journal of Biological Chemistry·Koustubh PandaDennis J Stuehr
Nov 28, 2006·The Journal of Biological Chemistry·Zhi-Qiang WangDennis J Stuehr
Feb 18, 2004·The Journal of Biological Chemistry·Koustubh PandaDennis J Stuehr
Aug 31, 2020·Redox Biology·Kulwant S AulakRaed A Dweik
May 9, 2021·Molecular and Cellular Endocrinology·Virginia DelliKonstantina Chachlaki
Jan 5, 2011·Journal of Inorganic Biochemistry·Jérôme Santolini

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