Purification and characterization of S-adenosylhomocysteine hydrolase from mouse mastocytoma P-815 cells. Evidence for cell cycle-specific fluctuation of the enzyme activity.
S-Adenosylhomocysteine hydrolase [EC 3.3.1.1] was purified to electrophoretic homogeneity from mastocytoma P-815 cells. The purified enzyme had a molecular weight of 190,000, as estimated by Sephadex G-200 chromatography, and a monomer molecular weight of 45,000, as determined by polyacrylamide gel electrophoresis in the presence of SDS. The Km value for adenosine was 0.29 microM and the Vmax value 4.5 mumol S-adenosylhomocysteine X min-1 X mg-1 in the synthetic reaction, while the Km value for S-adenosylhomocysteine was 0.77 microM and the Vmax 0.48 mumol adenosine X min-1 X mg-1 in the hydrolytic reaction. The purified enzyme also had one binding site for adenosine (KD = 2.61 X 10(-7) M) and one for cAMP (KD = 1.6 X 10(-7) M). Using rabbit antiserum raised against the purified enzyme, it was shown that the enzyme activity and enzyme synthesis fluctuated during the cell cycle of mastocytoma cells, reaching the maximum levels as the cells changed from the G1/S phase to the G2 phase.
Ichikawa
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Sato
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Tomita
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D000241:Adenosine / Q000378:metabolism; D043383:Adenosylhomocysteinase; D000818:Animals; D002453:Cell Cycle; D002460:Cell Line; D002850:Chromatography, Gel; D000242:Cyclic AMP / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D006867:Hydrolases / Q000302:isolation & purification / Q000378:metabolism*; D005779:Immunodiffusion; D007399:Interphase; D007526:Isoelectric Point; D007700:Kinetics; D012515:Mast-Cell Sarcoma / Q000201:enzymology*; D051379:Mice; D008970:Molecular Weight
D000242:Cyclic AMP; D006867:Hydrolases; D043383:Adenosylhomocysteinase; D000241:Adenosine