Direct interaction of endothelial nitric-oxide synthase and caveolin-1 inhibits synthase activity

Hong Ju, Rong Zou, Virginia J. Venema, Richard C. Venema

Research output: Contribution to journalArticle

480 Citations (Scopus)

Abstract

Endothelial nitric-oxide synthase (eNOS) and caveolind are associated within endothelial plasmalemmal caveolae. It is not known, however, whether eNOS and caveolin-1 interact directly or indirectly or whether the interaction affects eNOS activity. To answer these questions, we have cloned the bovine caveolin-1 cDNA and have investigated the eNOS-caveolin-1 interaction in an in vitro binding assay system using glutathione S- transferase (GST)-caveolin-1 fusion proteins and baculovirus-expressed bovine eNOS. We have also mapped the domains involved in the interaction using an in vivo yeast two-hybrid system. Results obtained using both in vitro and in vivo protein interaction assays show that both N- and C-terminal cytosolic domains of caveolin-1 interact directly with the eNOS oxygenase domain. Interaction of eNOS with GST-caveolin-1 fusion proteins significantly inhibits enzyme catalytic activity. A synthetic peptide corresponding to caveolin-1 residues 82-101 also potently and reversibly inhibits eNOS activity by interfering with the interaction of the enzyme with Ca2+/calmodulin (CaM). Regulation of eNOS in endothelial cells, therefore, may involve not only positive allosteric regulation by Ca/2+/CaM, but also negative allosteric regulation by caveolin-1.

Original languageEnglish (US)
Pages (from-to)18522-18525
Number of pages4
JournalJournal of Biological Chemistry
Volume272
Issue number30
DOIs
StatePublished - Aug 12 1997

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Caveolin 1
Nitric Oxide Synthase Type III
Allosteric Regulation
Calmodulin
Glutathione Transferase
Assays
Fusion reactions
Caveolae
Oxygenases
Two-Hybrid System Techniques
Proteins
Baculoviridae
Endothelial cells
Enzymes
Hybrid systems
Yeast
Catalyst activity
Endothelial Cells
Complementary DNA
Peptides

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

Cite this

Direct interaction of endothelial nitric-oxide synthase and caveolin-1 inhibits synthase activity. / Ju, Hong; Zou, Rong; Venema, Virginia J.; Venema, Richard C.

In: Journal of Biological Chemistry, Vol. 272, No. 30, 12.08.1997, p. 18522-18525.

Research output: Contribution to journalArticle

Ju, Hong ; Zou, Rong ; Venema, Virginia J. ; Venema, Richard C. / Direct interaction of endothelial nitric-oxide synthase and caveolin-1 inhibits synthase activity. In: Journal of Biological Chemistry. 1997 ; Vol. 272, No. 30. pp. 18522-18525.
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