In vitro generation of an active calmodulin-independent phosphodiesterase from brain calmodulin-dependent phosphodiesterase (PDE1A2) by m-calpain.
In the present study we have shown that bovine brain 60-kDa calmodulin-dependent cyclic nucleotide phosphodiesterase isozyme (CaMPDE - PDE1A2) is proteolyzed by a Ca2+-dependent cysteine protease, m-calpain. The proteolysis of PDE1A2 by m-calpain results in its conversion to a totally calmodulin (CaM)-independent form accompanied by degradation of PDE1A2 into a 45-kDa catalytic fragment and a 15-kDa fragment. The activity of PDE1A2 is unaffected by the presence or absence of CaM during cleavage, suggesting that the interaction between CaM and PDE1A2 does not alter substrate recognition by calpain. Furthermore, we provide evidence, based on the studies of CaM overlay and phosphorylation, that the cleavage site is not present either in the CaM-binding domain or phosphorylation site. N-terminal sequence analysis of the 45-kDa fragment indicated that cleavage occurs between residues 126Gln and 127Ala, and eliminates the CaM-dependent activity of carboxy termini PDE1A2. The present findings suggest that limited proteolysis in the brain through calpains could be an alternate mechanism for activating CaMPDE(s) and for regulating intracellular levels of cAMP.
College of Medicine, University of Saskatchewan, Saskatoon, S7N 4H4, Canada.
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D015105:3',5'-Cyclic-AMP Phosphodiesterases / Q000378:metabolism*; D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D001921:Brain / Q000201:enzymology*; D002118:Calcium / Q000378:metabolism; D002147:Calmodulin / Q000378:metabolism / Q000502:physiology; D002154:Calpain / Q000378:metabolism*; D002417:Cattle; D054677:Cyclic Nucleotide Phosphodiesterases, Type 1; D004789:Enzyme Activation; D006868:Hydrolysis; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010446:Peptide Fragments / Q000378:metabolism; D010727:Phosphoric Diester Hydrolases / Q000096:biosynthesis* / Q000737:chemistry / Q000378:metabolism*; D010766:Phosphorylation; D011485:Protein Binding
D002147:Calmodulin; D010446:Peptide Fragments; D010727:Phosphoric Diester Hydrolases; D015105:3',5'-Cyclic-AMP Phosphodiesterases; D054677:Cyclic Nucleotide Phosphodiesterases, Type 1; D002154:Calpain; D002118:Calcium