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Endothelial nitric oxide synthase. N-terminal myristoylation determines subcellular localization
1Cardiovascular Division, Brigham and Women's Hospital, Harvard Medical School, Boston, Massachusetts 02115.
The Journal of Biological Chemistry
|April 25, 1993
Summary
Endothelial nitric oxide synthase (eNOS) is localized to cell membranes via N-terminal myristoylation. This modification is crucial for eNOS function and may regulate nitric oxide production in endothelial cells.
Area of Science:
- Biochemistry
- Molecular Biology
- Cell Biology
Background:
- Nitric oxide synthases (NOS) produce nitric oxide (NO), a vital signaling molecule.
- Endothelial NOS (eNOS) is a distinct isoform localized to the particulate fraction, unlike soluble NOS isoforms.
- eNOS lacks transmembrane regions but has an N-terminal myristoylation motif.
Purpose of the Study:
- To investigate the role of N-terminal myristoylation in the subcellular localization and function of eNOS.
- To determine if myristoylation targets eNOS to specific cellular compartments.
Main Methods:
- Oligonucleotide-directed mutagenesis of the eNOS cDNA myristoylation site.
- Transient transfection of COS-7 cells with wild-type and mutant eNOS cDNA.
- Analysis of enzyme activity and immunoprecipitation of NO synthase.
- Biosynthetic labeling with [3H]myristate.
Main Results:
- Wild-type eNOS localized to the particulate fraction in transfected cells.
- Mutant eNOS lacking the myristoylation site localized exclusively to the cytosol.
- Myristoylation was confirmed by [3H]myristate labeling of wild-type eNOS in the particulate fraction.
Conclusions:
- N-terminal myristoylation is essential for targeting eNOS to the particulate fraction.
- This post-translational modification may regulate the biological activity of endothelium-derived NO.