Calmodulin-induced Conformational Control and Allostery Underlying Neuronal Nitric Oxide Synthase Activation

J Mol Biol. 2018 Mar 30;430(7):935-947. doi: 10.1016/j.jmb.2018.02.003. Epub 2018 Feb 17.

Abstract

Nitric oxide synthase (NOS) is the primary generator of nitric oxide signals controlling diverse physiological processes such as neurotransmission and vasodilation. NOS activation is contingent on Ca2+/calmodulin binding at a linker between its oxygenase and reductase domains to induce large conformational changes that orchestrate inter-domain electron transfer. However, the structural dynamics underlying activation of full-length NOS remain ambiguous. Employing hydrogen-deuterium exchange mass spectrometry, we reveal mechanisms underlying neuronal NOS activation by calmodulin and regulation by phosphorylation. We demonstrate that calmodulin binding orders the junction between reductase and oxygenase domains, exposes the FMN subdomain, and elicits a more dynamic oxygenase active site. Furthermore, we demonstrate that phosphorylation partially mimics calmodulin activation to modulate neuronal NOS activity via long-range allostery. Calmodulin binding and phosphorylation ultimately promote a more dynamic holoenzyme while coordinating inter-domain communication and electron transfer.

Keywords: allosteric communication; hydrogen–deuterium exchange; mass spectrometry; nitric oxide signaling.

Publication types

  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Allosteric Regulation
  • Calmodulin / metabolism*
  • Catalytic Domain
  • Deuterium Exchange Measurement
  • Enzyme Activation
  • Humans
  • Nitric Oxide Synthase Type I / chemistry*
  • Nitric Oxide Synthase Type I / metabolism*
  • Protein Conformation

Substances

  • Calmodulin
  • Nitric Oxide Synthase Type I