Purification and properties of the cGMP-inhibited cAMP phosphodiesterase from bovine aortic smooth muscle

Rascón, Ana; Lindgren, S.; Stavenow, Lars; Belfrage, Per; Andersson, Karl‐Erik; Manganiello, Vincent C.; Degerman, Eva

doi:10.1016/0167-4889(92)90038-d

Cited by 47 publications

(20 citation statements)

References 23 publications

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“…2), consistent with monomeric sizes of 130 -135 kDa for PDE3 isoforms identified in adipocyte microsomal and cardiac sarcoplasmic reticulum fractions (30,31). PDE3 isoforms of ϳ110 kDa and less have been purified from cytosolic fractions of human platelets, bovine ventricular myocardium, and bovine aortic smooth muscle (25,(32)(33)(34). The N-terminal hydrophobic regions, which contain several predicted transmembrane segments, are likely to be important in membrane association of PDE3s, since PDE3B and PDE3A recombinants (both full-length and truncated forms, which contain portions of the hydrophobic region) were found predominantly in particulate fractions of Sf9 cells, whereas recombinants containing the C-terminal catalytic core but lacking the hydrophobic sequences were predominantly cytosolic (21,27).…”

Section: Tissue-specific Expression Structure and Subcellularmentioning

confidence: 83%

Structure, Localization, and Regulation of cGMP-inhibited Phosphodiesterase (PDE3)

Degerman

Belfrage

Manganiello

1997

Journal of Biological Chemistry

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405

367

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Section: Tissue-specific Expression Structure and Subcellularmentioning

confidence: 83%

Structure, Localization, and Regulation of cGMP-inhibited Phosphodiesterase (PDE3)

Degerman

Belfrage

Manganiello

1997

Journal of Biological Chemistry

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405

367

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“…Type III PDEs (PDE3s), commonly referred to as cGMPinhibited cyclic nucleotide phosphodiesterases ( 11), have been isolated from a number of tissues, including adipose tissue ( 12), platelets (13), myocardium (14), vascular (15,16) and tracheo-bronchial (17,18) smooth muscle preparations, liver (19,20), placenta (21), and lymphocytes (22). PDE3s are characterized by a high affinity for cAMP and cGMP, with the V,,.…”

Section: Introductionmentioning

confidence: 99%

Distinctive anatomical patterns of gene expression for cGMP-inhibited cyclic nucleotide phosphodiesterases.

et al. 1995

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Type III cGMP-inhibited phosphodiesterases (PDE3s) play important roles in hormonal regulation of lipolysis, platelet aggregation, myocardial contractility, and smooth muscle relaxation. We have recently characterized two PDE3 subtypes (PDE3A and PDE3B) as products of distinct but related genes. To elucidate their biological roles, in this study we compare cellular patterns of gene expression for these two enzymes during rat embryonic and postnatal development using in situ hybridization. PDE3A mRNA is abundant in adipose tissue and is also expressed in hepatocytes throughout development. This mRNA is also highly abundant in embryonic neuroepithelium including the neural retina, but expression is greatly reduced in the mature nervous system. Finally, PDE3A mRNA is localized in spermatocytes and renal collecting duct epithelium in adult rats.PDE3B mRNA is highly expressed in the cardiovascular system, including myocardium and arterial and venous smooth muscle, throughout development. It is also abundant in bronchial, genitourinary and gastrointestinal smooth muscle and epithelium, megakaryocytes, and oocytes. PDE3B mRNA demonstrates a complex, developmentally regulated pattern of gene expression in the central nervous system. In summary, the two different PDE3s show distinctive tissue-specific patterns of gene expression suggesting that PDE3A is involved in hormonal regulation of lipolysis and glycogenolysis, while regulation of myocardial and smooth muscle contractility appears to be a function of PDE3B. In addition, the present findings suggest previously unsuspected roles for these enzymes in gametogenesis and neural development. (J. Clin. Invest. 1995Invest. . 95:1528Invest. -1538

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“…Studies of several PDEs have suggested that the isolated catalytic domain alone is sufficient for catalytic activity. However, the importance of dimerization to the catalytic function of the PDE remained in question (31,32,34,35). An amino-terminal truncation mutant of PDE1 was CaMindependent but maintained catalytic properties (K m and V max ) that were similar to those of wild type.…”

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confidence: 99%

Expression of an Active, Monomeric Catalytic Domain of the cGMP-binding cGMP-specific Phosphodiesterase (PDE5)

Fink¹,

Francis²,

Beasley

et al. 1999

Journal of Biological Chemistry

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Phosphodiesterases (PDEs) comprise a superfamily of phosphohydrolases that degrade 3,5-cyclic nucleotides. All known mammalian PDEs are dimeric, but the functional significance of dimerization is unknown. A deletion mutant of cGMP-binding cGMP-specific PDE (PDE5), encoding the 357 carboxyl-terminal amino acids including the catalytic domain, has been generated, expressed, and purified. The K m of the catalytic fragment for cGMP (5.5 ؎ 0.51 M) compares well with those of the native bovine lung PDE5 (5.6 M) and full-length wild type recombinant PDE5 (2 ؎ 0.4 M). The catalytic fragment and full-length PDE5 have similar IC 50 values for the inhibitors 3-isobutyl-1-methylxanthine (20 M) and sildenafil (Viagra TM )(4 nM). Based on measured values for Stokes radius (29 Å) and sedimentation coefficient (2.9 S), the PDE5 catalytic fragment has a calculated molecular mass of 35 kDa, which agrees well with that predicted by amino acid content (43.3 kDa) and with that estimated using SDS-polyacrylamide gel electrophoresis (39 kDa). The combined data indicate that the recombinant PDE5 catalytic fragment is monomeric, and retains the essential catalytic features of the dimeric, full-length enzyme. Therefore, the catalytic activity of PDE5 holoenzyme requires neither interaction between the catalytic and regulatory domains nor interactions between subunits of the dimer.Intracellular levels of cyclic nucleotide are determined both by their rates of formation by adenylyl cyclases and guanylyl cyclases and their rates of breakdown by cyclic nucleotide phosphodiesterases (PDEs) 1 (1). Both the cyclases and the PDEs are highly regulated (2-8). Mammalian PDEs form a superfamily of enzymes comprised of 10 families of PDEs which differ in amino acid sequences (2, 3, 9 -11), substrate specificities, inhibitor sensitivities, modes of regulation, and tissue distribution. PDEs are chimeric proteins (12-15) which contain specific domains that provide for cyclic nucleotide hydrolysis, interaction with allosteric/regulatory molecules, autoinhibition, subcellular localization, and perhaps for other functions as well (12,16,17). For mammalian PDEs, the regulatory domains appear to reside mainly in the amino-terminal portions of the proteins. The carboxyl-terminal portion of each PDE contains a highly conserved region of ϳ250 amino acids (4), which comprises the catalytic domain.The cGMP-binding cGMP-specific PDE (PDE5 or cG-BPDE), which is the focus of this study, is abundant in lung, platelets, and vascular smooth muscle (18 -23). It is potently inhibited by sildenafil (Viagra TM ), which has proven to be therapeutically efficacious in the treatment of male erectile dysfunction (24). The PDE5 is highly specific for cGMP, both in its catalytic site, and in the two cGMP-binding allosteric sites within the aminoterminal half of the protein (14, 25). The regulation of the PDE5 catalytic activity is not well understood, although evidence has been presented that catalytic function is influenced by elements within the regulatory domain (26,27). Cycl...

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Purification and properties of the cGMP-inhibited cAMP phosphodiesterase from bovine aortic smooth muscle

Cited by 47 publications

References 23 publications

Structure, Localization, and Regulation of cGMP-inhibited Phosphodiesterase (PDE3)

Structure, Localization, and Regulation of cGMP-inhibited Phosphodiesterase (PDE3)

Distinctive anatomical patterns of gene expression for cGMP-inhibited cyclic nucleotide phosphodiesterases.

Expression of an Active, Monomeric Catalytic Domain of the cGMP-binding cGMP-specific Phosphodiesterase (PDE5)

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