RRC ID |
58226
|
Author |
Adachi H, Iida S, Oguchi S, Ohshima H, Suzuki H, Nagasaki K, Kawasaki H, Sugimura T, Esumi H.
|
Title |
Molecular cloning of a cDNA encoding an inducible calmodulin-dependent nitric-oxide synthase from rat liver and its expression in COS 1 cells.
|
Journal |
Eur J Biochem
|
Abstract |
Calmodulin-dependent nitric-oxide synthase, with an apparent molecular mass of 125 kDa, was induced in the liver of rats treated with Propionibacterium acnes and Escherichia coli lipopolysaccharide. Clones were isolated from a cDNA library obtained from induced rat liver using oligonucleotide probes which were synthesized based on the amino acid sequences of peptides of the purified enzyme. Four overlapping cDNA clones for a 3.8-kbp region were isolated and the nucleotide sequences were determined. These clones encompassed an open-reading frame of 3441 bases encoding 1147 amino acids. The deduced amino acid sequence of the cDNA suggested that the protein contains binding sites for NADPH, FAD and FMN. The structure of the possible calmodulin-binding site, consisting of a strongly hydrophobic region surrounded by basic amino acids, is present. The full-length cDNA was expressed in COS 1 cells under the control of a cytomegalovirus promoter and the expressed enzyme was found to be a calmodulin-dependent nitric-oxide synthase. A structural comparison suggested that the liver nitric-oxide synthase is the same as the macrophage enzyme. Northern-blot analysis showed that the mRNA in the liver is approximately 4.2 kb long and is induced transcriptionally by treatment with P. acnes and lipopolysaccharide.
|
Volume |
217(1)
|
Pages |
37-43
|
Published |
1993-10-1
|
DOI |
10.1111/j.1432-1033.1993.tb18215.x
|
PMID |
7693462
|
MeSH |
Amino Acid Oxidoreductases / biosynthesis
Amino Acid Oxidoreductases / genetics*
Amino Acid Sequence
Animals
Base Sequence
Calmodulin / pharmacology*
Cell Line
Chlorocebus aethiops
Cloning, Molecular*
DNA Restriction Enzymes
DNA, Complementary / chemistry
DNA, Complementary / genetics*
Enzyme Induction / drug effects
Gene Expression*
Kidney
Liver / enzymology*
Male
Molecular Sequence Data
Nitric Oxide Synthase
Rats
Rats, Sprague-Dawley
Transfection
|
Times Cited |
64
|
Resource |
DNA material |
Inducible NOS cDNA (RDB01385) |