We have produced a polyclonal antibody that specifically recognizes cGMP-binding cGMP-specific phosphodiesterase (PDES). The antibody was raised in rabbit using as immunogen a fusion protein, in which glutathione S-transferase was coupled to a 171 amino acid polypeptide of the N-terminal region of bovine PDE5. The antibody is able to immunoprecipitate PDES activity from mouse tissues and neuroblastoma extracts while it has no effect on all other PDE isoforms present in the extracts. PDES activity recovered in the immunoprecipitates retains its sensitivity to specific inhibitors such as zaprinast (IC50 = 0.6 muM) and sildenafil (IC50 = 3.5 nM), Bands of the expected molecular mass were revealed when solubilized immunoprecipitates were analysed in Western blots. The antibody selectively stained cerebellar Purkinje neurones, which are known to express high levels of PDES mRNA. Western blot analysis of mouse tissues revealed the highest expression signal in mouse lung, followed by heart and cerebellum, while a lower signal was evident in brain, kidney and a very low signal was present in the liver. In the hybrid neuroblastoma-glioma NG108-15 cells the antibody revealed a high PDE5 induction after dibutyryl-cAMP treatment. (C) 2001 Elsevier Science B,V, All rights reserved.

Expression of cGMP-binding cGMP-specific phosphodiesterase (PDE5) in mouse tissues and cell lines using an antibody against the enzyme amino-terminal domain / Daniela, Giordano; DE STEFANO, Maria Egle; Gennaro, Citro; Anna, Modica; Giorgi, Mauro. - In: BIOCHIMICA ET BIOPHYSICA ACTA-MOLECULAR CELL RESEARCH. - ISSN 0167-4889. - STAMPA. - 1539:1-2(2001), pp. 16-27. [10.1016/s0167-4889(01)00086-6]

Expression of cGMP-binding cGMP-specific phosphodiesterase (PDE5) in mouse tissues and cell lines using an antibody against the enzyme amino-terminal domain

DE STEFANO, Maria Egle;GIORGI, MAURO
2001

Abstract

We have produced a polyclonal antibody that specifically recognizes cGMP-binding cGMP-specific phosphodiesterase (PDES). The antibody was raised in rabbit using as immunogen a fusion protein, in which glutathione S-transferase was coupled to a 171 amino acid polypeptide of the N-terminal region of bovine PDE5. The antibody is able to immunoprecipitate PDES activity from mouse tissues and neuroblastoma extracts while it has no effect on all other PDE isoforms present in the extracts. PDES activity recovered in the immunoprecipitates retains its sensitivity to specific inhibitors such as zaprinast (IC50 = 0.6 muM) and sildenafil (IC50 = 3.5 nM), Bands of the expected molecular mass were revealed when solubilized immunoprecipitates were analysed in Western blots. The antibody selectively stained cerebellar Purkinje neurones, which are known to express high levels of PDES mRNA. Western blot analysis of mouse tissues revealed the highest expression signal in mouse lung, followed by heart and cerebellum, while a lower signal was evident in brain, kidney and a very low signal was present in the liver. In the hybrid neuroblastoma-glioma NG108-15 cells the antibody revealed a high PDE5 induction after dibutyryl-cAMP treatment. (C) 2001 Elsevier Science B,V, All rights reserved.
cyclic guanosine monophosphate; glutathione s transferase fusion protein; glutathione s-transferase fusion protein; murine neuroblastoma; phosphodiesterase; phosphodiesterase-5 antibody
01 Pubblicazione su rivista::01a Articolo in rivista
Expression of cGMP-binding cGMP-specific phosphodiesterase (PDE5) in mouse tissues and cell lines using an antibody against the enzyme amino-terminal domain / Daniela, Giordano; DE STEFANO, Maria Egle; Gennaro, Citro; Anna, Modica; Giorgi, Mauro. - In: BIOCHIMICA ET BIOPHYSICA ACTA-MOLECULAR CELL RESEARCH. - ISSN 0167-4889. - STAMPA. - 1539:1-2(2001), pp. 16-27. [10.1016/s0167-4889(01)00086-6]
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Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/11573/42971
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