The extracellular cyclic nucleotide phosphodiesterase of Dictyostelium discoideum. Purification and characterization.
Two forms of the extracellular cyclic nucleotide phosphodiesterase (EC 3.1.4.17) of Dictyostelium discoideum have been purified. One species has a molecular weight of about 55,000 measured by gel filtration and has been purified to apparent homogeneity. This monomeric form of the enzyme can be resolved by isoelectrofocusing into 4 bands with isoelectric points between 7.5 and 9. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed two bands with molecular weights of 50,000 and 48,000. The second form of the enzyme has been partially purified and has a higher molecular weight (150,000-200,000) and an isoelectric point of about 5. The high molecular weight form of the enzyme is converted to the low molecular weight monomer by isoelectrofocusing in the presence of 6 M urea. Under these conditions the isoelectric point is shifted to that of the monomeric form of the enzyme. The two species are immunologically indistinguishable after urea treatment, and react identically with the cyclic nucleotide phosphodiesterase inhibitor (see the accompanying paper: Franke, J., and Kessin, R. H. (1981) J. Biol. Chem. 256, 7628-7637). The two forms have the same Km and show equal sensitivity to a number of ions, chelators, and low molecular weight cyclic nucleotide phosphodiesterase inhibitors. Tryptic maps of the purified monomeric enzyme and the urea dissociated high molecular weight form revealed no differences.
D015105:3',5'-Cyclic-AMP Phosphodiesterases / Q000302:isolation & purification / Q000378:metabolism*; D002413:Cations, Divalent; D004023:Dictyostelium / Q000201:enzymology*; D007527:Isoenzymes / Q000302:isolation & purification / Q000378:metabolism; D007700:Kinetics; D008970:Molecular Weight; D010446:Peptide Fragments / Q000032:analysis; D013438:Sulfhydryl Compounds / Q000494:pharmacology; D014357:Trypsin; D014508:Urea / Q000494:pharmacology
D002413:Cations, Divalent; D007527:Isoenzymes; D010446:Peptide Fragments; D013438:Sulfhydryl Compounds; D014508:Urea; D015105:3',5'-Cyclic-AMP Phosphodiesterases; D014357:Trypsin