In vitro PKA phosphorylation-mediated human PDE4A4 activation |
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Authors: | Laliberté France Liu Susana Gorseth Elise Bobechko Brian Bartlett Adrienne Lario Paula Gresser Michael J Huang Zheng |
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Institution: | Department of Biochemistry and Molecular Biology, Merck Frosst Centre for Therapeutical Research, P.O. Box 1005, Pointe Claire, H9R4P8, Dorval, QC, Canada. |
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Abstract: | The PDE4 catalytic machinery comprises, in part, two divalent cations in a binuclear motif. Here we report that PDE4A4 expressed in Sf9 cells exhibits a biphasic Mg(2+) dose-response (EC(50) of 0.15 and >10 mM) in catalyzing cAMP hydrolysis. In vitro phosphorylation of PDE4A4 by the PKA-catalytic subunit increases the enzyme's sensitivity to Mg(2+), leading to 4-fold increased cAMP hydrolysis without affecting its K(m). The phosphorylation also increases the potencies of (R)- and (S)-rolipram without affecting CDP-840 and SB-207499. The results support that modulating the cofactor binding affinity of PDE4 represents a mechanism for regulating its activity. |
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