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Regulation of Nitric Oxide-Responsive Recombinant Soluble Guanylyl Cyclase by Calcium
- Source :
- Biochemistry. 38:6441-6448
- Publication Year :
- 1999
- Publisher :
- American Chemical Society (ACS), 1999.
-
Abstract
- Calcium (Ca2+) and cyclic GMP (cGMP) subserve antagonistic functions that are reflected in their coordinated reciprocal regulation in physiological systems. However, molecular mechanisms by which Ca2+ regulates cGMP-dependent signaling remain incompletely defined. In this study, the inhibition of recombinant nitric oxide (NO)-stimulated soluble guanylyl cyclase (SGC) by Ca2+ was demonstrated. The alpha- and beta-subunits of recombinant rat SGC were heterologously coexpressed in HEK 293 cells which do not express NO synthase, whose Ca2+-stimulated activity can confound the effects of that cation on SGC. Ca2+ inhibited basal and NO-stimulated SGC in a concentration- and guanine nucleotide-dependent fashion. This cation inhibited SGC in crude cell extracts and immunopurified preparations. Ca2+ lowered both the Vmax and Km of SGC via an uncompetitive mechanism through direct interaction with the enzyme. In intact HEK 293 cells, increases in the intracellular Ca2+ concentration induced by ionomycin, a Ca2+ ionophore, and thapsigargin, which releases intracellular stores of that cation, inhibited NO-stimulated intracellular cGMP accumulation. Similarly, carbachol-induced elevation of the intracellular Ca2+ concentration inhibited NO-stimulated intracellular cGMP accumulation in HEK 293 cells. These data demonstrate that SGC behaves as a sensitive Ca2+ detector that may play a central role in coordinating the reciprocal regulation of Ca2+- and cGMP-dependent signaling mechanisms.
- Subjects :
- Cell Extracts
Intracellular Fluid
inorganic chemicals
GUCY1B3
Thapsigargin
chemistry.chemical_element
Calcium
Kidney
Nitric Oxide
Binding, Competitive
Biochemistry
Cell Line
Nitric oxide
chemistry.chemical_compound
Animals
Humans
Enzyme Inhibitors
Cyclic GMP
HEK 293 cells
Guanine Nucleotides
Recombinant Proteins
Rats
Cell biology
Solubility
chemistry
Guanylate Cyclase
Ionomycin
cardiovascular system
Guanosine Triphosphate
Soluble guanylyl cyclase
Intracellular
Subjects
Details
- ISSN :
- 15204995 and 00062960
- Volume :
- 38
- Database :
- OpenAIRE
- Journal :
- Biochemistry
- Accession number :
- edsair.doi.dedup.....27c99866332a56a83dc88cdb364b4554