Inhibition of human phosphodiesterase 4A expressed in yeast cell GL62 by theophylline, rolipram, and acetamide-45.

Kai Wang, Ji-Qiang Chen, Zhong Chen, Jun-Chun Chen

Respiratory Drugs Research Laboratory of State Drugs Administration of China,Medical School of Zhejiang University, Hangzhou 310031, China.

Acta pharmacologica Sinica 2002 Nov

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To study the inductive expression of human phosphodiesterase 4A (hPDE4A) in yeast cell GL62 and investigate the inhibitory effects of theophylline, rolipram, and acetamide-45 on PDE4A activity of the expressed product in yeast cell GL62. Yeast cell GL62 were transfected with human PDE4A gene cloned in the expression plasmid p138NB. Expression was induced by adding CuSO4 to a final concentration of 150 micromol/L, and the expressed product was extracted. The activity of PDE4A was detected by HPLC. Yeast cell GL62 expressed a certain protein at CuSO4 150 micromol/L, the size of the expressed product was between 62 kDa and 83 kDa, the activity of PDE4A of the expressed product at 3 h was in maximum (188 23) micromol/g/min, and the Km was (17.7 2.6) micromol/L. Theophylline, rolipram, and acetamide-45 could inhibit the activity of PDE4A extracted from yeast cell GL62. The IC50 (95 % confidence limits) of theophylline, rolipram, and acetamide-45 were 1642 (989-2727), 4.58 (3.45-6.08), and 275 (170-444) micromol/L respectively. PDE4A expressed in yeast cell GL62 is biologically active. Theophylline, rolipram, and acetamide-45 can inhibit the PDE4A activity. The expressed product in yeast cell GL62 may be used in the research work of PDE4 and its inhibitors.

Citation

Kai Wang, Ji-Qiang Chen, Zhong Chen, Jun-Chun Chen. Inhibition of human phosphodiesterase 4A expressed in yeast cell GL62 by theophylline, rolipram, and acetamide-45. Acta pharmacologica Sinica. 2002 Nov;23(11):1013-7