Identification and stabilization of large molecular weight PDE-IVs from U937 cells

Biochem Biophys Res Commun. 1993 Dec 30;197(3):1126-31. doi: 10.1006/bbrc.1993.2594.

Abstract

Cytosolic cyclic nucleotide phosphodiesterases (PDEs) from human (promonocytic) U937 cells were rapidly resolved by DEAE-Sepharose CL-6B anion exchange chromatography into two major peaks of cAMP-specific activity possessing average Kms of 1.70 microM (Peak 1) and 1.65 microM (Peak 2). Both peaks were predominantly PDE-IV, but possessed molecular weights higher than those generally reported for partially purified PDE-IVs. Storage of Peak 2 for 24 h at 4 degrees C resulted in a doubling of its Vmax and an apparent decrease in its molecular weight. Activation of Peak 2 PDE-IV was prevented when the sodium acetate concentration in its buffer was reduced by dilution immediately following isolation. Although the relevance of this activation to cellular regulation of PDE-IV is undefined, the isolation and stabilization of PDE-IV in its large molecular weight form will be critical to future investigations of PDE-IV regulation.

MeSH terms

  • 3',5'-Cyclic-AMP Phosphodiesterases / chemistry
  • 3',5'-Cyclic-AMP Phosphodiesterases / isolation & purification*
  • 3',5'-Cyclic-AMP Phosphodiesterases / metabolism
  • Cell Line
  • Chromatography, Ion Exchange
  • Cytosol / enzymology
  • Enzyme Stability
  • Humans
  • Kinetics
  • Molecular Weight
  • Substrate Specificity
  • Time Factors
  • Tumor Cells, Cultured

Substances

  • 3',5'-Cyclic-AMP Phosphodiesterases