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Purification of a Ca 2+ /calmodulin‐dependent nitric oxide synthase from porcine cerebellum
Author(s) -
Mayer Bernd,
John Mathias,
Böhme Eycke
Publication year - 1990
Publication title -
febs letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.593
H-Index - 257
eISSN - 1873-3468
pISSN - 0014-5793
DOI - 10.1016/0014-5793(90)80848-d
Subject(s) - tetrahydrobiopterin , nitric oxide synthase , biochemistry , cytosol , calmodulin , ammonium sulfate precipitation , enzyme , cofactor , atp synthase , chemistry , sepharose , soluble guanylyl cyclase , affinity chromatography , nitric oxide , microbiology and biotechnology , biology , guanylate cyclase , organic chemistry , size exclusion chromatography
L‐Arginine‐derived nitric oxide acts as an inter‐ and intracellular signal molecule with cytosolic guanylyl cyclase as the effector system. Two NO synthase isoenzymes are postulated: a cytokine‐inducible enzyme in macrophages and a constitutive, Ca 2+ ‐regulated enzyme in various other cells. An NO synthase was isolated from porcine cerebellum by ammonium sulfate precipitation and affinity chromatography on 2',5'‐ADP‐Sepharose. The enzyme was identified as an NO synthase with a specific NO‐chemiluminescence method and with purified cytosolic guanylyl cyclase as an NO‐sensitive detection system. The purified NO synthase was, besides Ca 2+ /calmodulin and NADPH, largely dependent on tetrahydrobiopterin as a cofactor.