Subunit interactions of endothelial nitric-oxide synthase: Comparisons to the neuronal and inducible nitric-oxide synthase isoforms

Richard C Venema, Hong Ju, Kong Zou, James W. Ryan, Virginia J. Venema

Research output: Contribution to journalArticle

116 Scopus citations

Abstract

Endothelial nitric-oxide synthase (eNOS) is comprised of two identical subunits. Each subunit has a bidomain structure consisting of an N-terminal oxygenase domain containing heme and tetrahydrobiopterin (BH4) and a C- terminal reductase domain containing binding sites for FAD, FMN, and NADPH. Each subunit is also myristoylated and contains a calmodulin (CaM)-binding site located between the oxygenase and reductase domains. In this study, wild-type and mutant forms of eNOS have been expressed in a baculovirus system, and the quaternary structure of the purified enzymes has been analyzed by low temperature SDS-PAGE. eNOS dimer formation requires incorporation of the heme prosthetic group but does not require myristoylation or CaM or BH4 binding. In order to identify domains of eNOS involved in subunit interactions, we have also expressed eNOS oxygenase and reductase domain fusion proteins in a yeast two-hybrid system. Corresponding human neuronal NOS (nNOS) and murine inducible NOS (iNOS) fusion proteins have also been expressed. Comparative analysis of NOS domain interactions shows that subunit association of eNOS and nNOS involves not only head to head interactions of oxygenase domains but also tail to tail interactions of reductase domains and head to tail interactions between oxygenase and reductase domains. In contrast, iNOS subunit association involves only oxygenase domain interactions.

Original languageEnglish (US)
Pages (from-to)1276-1282
Number of pages7
JournalJournal of Biological Chemistry
Volume272
Issue number2
DOIs
StatePublished - Jan 25 1997

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

Fingerprint Dive into the research topics of 'Subunit interactions of endothelial nitric-oxide synthase: Comparisons to the neuronal and inducible nitric-oxide synthase isoforms'. Together they form a unique fingerprint.

  • Cite this