We developed a kinetic method for measuring the activities of adenosine deaminase (EC 3.5.4.4, adenosine aminohydrolase: ADA) isozymes, based on their different Km values for adenosine as substrate. ADA activity was measured using two different concentrations of substrate; high (20 mmol/l) and low (0.5 mmol/l) concentration solutions of adenosine.
This kinetic method dose not require gel filtration, and is thus a relatively simple and rapid procedure.
Serum ADA is separated by gel filtration into two types of isozymes which have been called ADAi (small form; 35,000 daltons and large form; 280,000 daltons) and ADA
2 (intermediate form; 100,000 daltons). ADA
2 shows markedly different kinetic properties from those of ADA
1; ADA
2 has a higher Km value for adenosine and a lower deamination activity for 2'-deoxyadenosine than ADA
1.
Analytical recoveries of ADA isozymes by the kinetic method varied from 93.3 to 116.3%. Results of the present method correlated well with those of the gel filtration method in high-performance liquid chromatography.
In clinical application, increased serum ADA
2 activity was observed in chronic liver diseases, such as chronic hepatitis, liver cirrhosis, and hepato-celluler carcinoma. However, serum ADA
1 activity did not increase significantly in only a few of many cases chronic liver disease. Studies on serial measurements of serum ADA isozymes in patients with liver disorders have demonstrated that the changes of ADA
1 activity are similar to those of aspartate aminotransferase and alanine aminotransferase activity, but the changes of ADA
2 activity differ.
These results suggest that measurement of serum ADA isozymes activities by the present method is useful for testing liver function.
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