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Acta Pharmacol Sin. 2002 Nov;23(11):1013-7.

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

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  • 1Respiratory Drugs Research Laboratory of State Drugs Administration of China,Medical School of Zhejiang University, Hangzhou 310031, China.

Abstract

AIM:

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.

METHODS:

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.

RESULTS:

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.

CONCLUSION:

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.

PMID:
12421478
[PubMed - indexed for MEDLINE]
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