title: Cyclic GMP-dependent protein kinase activity in rat pulmonary microvascular endothelial cells.
abstract: Cyclic GMP-dependent protein kinase (cGPK) activity was determined in rat pulmonary microvascular endothelial cells (RPMVEC) using cGMP-stimulated phosphorylation of BPDEtide and histone F2B substrates in the presence of PKI [peptide inhibitor of cAMP-dependent protein kinase (cAPK)]. RPMVEC cGPK activity was localized to the 100,000 x g cytosolic fraction. The EC50 for cGMP activation in the presence of PKI was 0.16 microM and H-89 inhibition under similar conditions showed an IC50 value of 0.16 microM. Anion-exchange chromatography of RPMVEC and rat lung cytosolic fractions showed separation of the cGMP-dependent from the cGMP-independent protein kinase activity and similar elution conductivities. Further, Western blots of RPMVEC active DEAE-Trisacryl fractions showed immunoreactivity using bovine Type I cGPK antiserum. Preliminary studies reveal six potential substrates phosphorylated by cGPK in RPMVEC. These studies describe an endothelial cell (EC) cGMP-receptor, cGPK, in addition to cGMP-activated (Type II) phosphodiesterase (PDE).
authors.affiliation: Dept. of Pharmacology, University of South Alabama College of Medicine, Mobile 36688.
authors.forename: A H
authors.initials: AH
authors.lastname: Diwan
authors.forename: W J
authors.initials: WJ
authors.lastname: Thompson
authors.forename: A K
authors.initials: AK
authors.lastname: Lee
authors.forename: S J
authors.initials: SJ
authors.lastname: Strada
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000838:Anions; D015153:Blotting, Western; D002852:Chromatography, Ion Exchange; D006152:Cyclic GMP / Q000494:pharmacology*; D003600:Cytosol / Q000201:enzymology; D004730:Endothelium, Vascular / Q000201:enzymology*; D008168:Lung / Q000098:blood supply*; D008833:Microcirculation / Q000201:enzymology; D008969:Molecular Sequence Data; D010766:Phosphorylation; D047428:Protein Kinase Inhibitors; D011494:Protein Kinases / Q000378:metabolism*; D051381:Rats; D012965:Sodium Chloride; D013379:Substrate Specificity
chemical_list: D000838:Anions; D047428:Protein Kinase Inhibitors; D012965:Sodium Chloride; D011494:Protein Kinases; D006152:Cyclic GMP