Complete release of adenosine deaminase from mouse lymphocytes stabilized by low-pH acetate.
Complete release of adenosine deaminase from mouse lymphocytes takes place when intact cells are stabilized by low-pH acetate buffer. Both the low pH and the acetate affect the enzyme extraction markedly. At pH 5.0 all the adenosine deaminase activity detectable in the whole cell homogenates is released into the acetate buffer in very few minutes, with a total amount of 2% protein being extracted. The complete extraction of the enzyme activity is never observed when, at pH 5.0, the acetate is replaced by glutamate, citrate, succinate or maleate and only 45% and 15% of the adenosine deaminase activity is extracted by the acetate at pH 6.0 and 7.0, respectively. The breakdown of adenosine by the enzyme activity extracted from the stabilized cells is due to deamination alone, since inosine is the only product of the catalyzed reaction and its formation is completely inhibited by coformycin, a selective inhibitor of adenosine deaminase. The enzyme extracted shows a specific activity 50-times higher than that found in the crude homogenates, and a substantial purification of the enzyme extracted is achieved by a single Sephadex G-100 gel filtration.
Senesi
S
S
Institute of Microbiology, University of Pisa, Italy.
Freer
G
G
Gasperini
M
M
Batoni
G
G
Campa
M
M
D000085:Acetates; D000243:Adenosine Deaminase / Q000378:metabolism*; D000818:Animals; D001402:B-Lymphocytes / Q000145:classification / Q000201:enzymology; D002021:Buffers; D002478:Cells, Cultured; D005260:Female; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008198:Lymph Nodes / Q000201:enzymology; D008214:Lymphocytes / Q000201:enzymology*; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D009700:Nucleoside Deaminases / Q000378:metabolism*; D013154:Spleen / Q000201:enzymology; D013601:T-Lymphocytes / Q000145:classification / Q000201:enzymology; D013950:Thymus Gland / Q000201:enzymology
D000085:Acetates; D002021:Buffers; D009700:Nucleoside Deaminases; D000243:Adenosine Deaminase