Adenosine deaminase isoenzymes of the opossum Didelphis virginiana: initial chromatographic and kinetic studies

Extracts of liver and spleen were used to isolate opossum adenosine deaminase isoenzymes (ADA, and ADA2) and to determine their activities with adenosine and 2′-deoxyadenosine as substrates. K_m values (μM) for adenosine and 2′-deoxyadenosine, respectively, as substrates for partially purified opossum liver adenosine deaminase isoenzymes were ADA,: 57 ± 7 vs. 26 ± 4 and ADA₂: 285 ± 25 vs. 580 ± 92. In crude spleen extract, ADA₂ activity was stable at 56°C during 40 min of incübation. ADA₁ activity declined in a linear fashion under the above conditions with an apparent T _{1 2} of 80 min. Sephadex G-150 column chromatography of crude spleen extract showed the apparent molecular weight of the ADA activity not inhibited by (±)-EHNA (i.e. ADA₂) to be 170 kDa; ADA activity fully inhibited by (±)-EHNA (i.e. ADA,) eluted in the fractions corresponding to a molecular weight of 35 kDa.