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Cloning and expression of cDNA for a human low-Km, rolipram-sensitive cyclic AMP phosphodiesterase
- Source :
- ResearcherID
- Publication Year :
- 1990
-
Abstract
- We have isolated cDNA clones representing cyclic AMP (cAMP)-specific phosphodiesterases (PDEases) from a human monocyte cDNA library. One cDNA clone (hPDE-1) defines a large open reading frame of ca. 2.1 kilobases, predicting a 686-amino-acid, ca. 77-kilodalton protein which contains significant homology to both rat brain and Drosophila cAMP PDEases, especially within an internal conserved domain of ca. 270 residues. Amino acid sequence divergence exists at the NH2 terminus and also within a 40- to 100-residue domain near the COOH-terminal end. hPDE-1 hybridizes to a major 4.8-kilobase mRNA transcript from both human monocytes and placenta. The coding region of hPDE-1 was engineered for expression in COS-1 cells, resulting in the overproduction of cAMP PDEase activity. The hPDE-1 recombinant gene product was identified as a low-Km cAMP phosphodiesterase on the basis of several biochemical properties including selective inhibition by the antidepressant drug rolipram. Known inhibitors of other PDEases (cGMP-specific PDEase, cGMP-inhibited PDEase) had little or no effect on the hPDE-1 recombinant gene product. Human genomic Southern blot analysis suggests that this enzyme is likely to be encoded by a single gene. The presence of the enzyme in monocytes may be important for cell function in inflammation. Rolipram sensitivity, coupled with homology to the Drosophila cAMP PDEase, which is required for learning and memory in flies, suggests an additional function for this enzyme in neurobiochemistry.
- Subjects :
- Phosphodiesterase Inhibitors
Immunoblotting
Molecular Sequence Data
Restriction Mapping
Gene Expression
Biology
Transfection
Cell Line
Gene product
Complementary DNA
Gene expression
medicine
Animals
Humans
Amino Acid Sequence
Cloning, Molecular
Molecular Biology
Rolipram
Cyclic nucleotide phosphodiesterase
Base Sequence
cDNA library
Phosphodiesterase
Cell Biology
DNA
Molecular biology
Pyrrolidinones
Recombinant Proteins
Isoenzymes
Open reading frame
Kinetics
Liver
3',5'-Cyclic-AMP Phosphodiesterases
medicine.drug
Research Article
Subjects
Details
- Language :
- English
- Database :
- OpenAIRE
- Journal :
- ResearcherID
- Accession number :
- edsair.doi.dedup.....705710f14b8d91169594ef0e135a9843