Structural basis for CA2+-independent activity of inducible nitric oxide synthase

J. Lee, J. T. Stull

Research output: Contribution to journalArticle

Abstract

Nitric oxide synthases (NOS) contain three subdomains, the N-terminal monooxygenase domain, the C-terminal reductase domain and the connecting canonical calmodulin(CaM)- binding domain. Neuronal NOS(nNOS) activity is Ca+/CaM-dependen, whereas inducible NOS (iNOS) activity is Ca2+independent presumably due to tightly bound CaM. To study the mechanism responsible for iNOS Ca2+-independent activity and nNOS Ca2+-dependent activity, a series of chimeric NOSs derived from nNOS and iNOS were transiently expressed in COS-7 cells. Ca2+-dependent NOS activities were measured in cell lysates in the presence of CaM. Results indicate that Ca2+independent activity ofiNOS results primarily from its monooxygenase domain and the canonical CaM-binding region acting in concert. However, neither the iNOS monooxygenase domain nor the canonical CaM-binding region alone confers Ca2+-independent NOS activity. Replacement of CaM-binding sequence of nNOS with that of iNOS does not confer Ca2+-independent NOS activity. Replacement of CaM-binding sequence of iNOS with that of nNOS changes the activity to Ca2+-dependent. In both cases, there is increased Ca2+sensitivity compared to nNOS. Additionally, the reductase domain of iNOS increases the Ca2+-sensitivity of a chimeric NOS containing nNOS CaM-binding sequence and the nNOS monoxygenase domain. The Ca2+-independent activity of iNOS appears to be mediated through different regions of the molecule, rather than the canonical CaM-binding region alone.

Original languageEnglish (US)
Pages (from-to)A1312
JournalFASEB Journal
Volume11
Issue number9
StatePublished - Dec 1 1997

ASJC Scopus subject areas

  • Biotechnology
  • Biochemistry
  • Molecular Biology
  • Genetics

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