A study of frog adenosine deaminases : purification and some properties

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Cook, Kenneth Steven
Ma, Pang-Fai
Issue Date
Thesis (M.S.)--Ball State University, 1983.
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Adenosine deaminase has been shown to consist of three molecular weight forms, A, B, and C. In higher mammals, the A and C forms are dominant while in lower mammals, the B and C forms are dominant. In this work, the B and C forms were isolated from the frog liver and several kinetic parameters were determined.Ammonium sulfate salt fractionation, starting at 40 percent and increased by 5 percent increments to 80 percent was used to separate the two forms. The B form adenosine deaminase was predominantly found in the 50 to 60 percent precipitate fractions while the C form was predominant in precipitate fractions containing more than 60 percent ammonium sulfate. The rechromatographed B and C forms were subjected to isoelectric focusing and thin layer electrophoresis. The B form separated into three activity bands while the C form separated into two activity bands`. Michaelis constant values were determined to be 4.61 X 10-5M and 2.00 X 10-5M for the B and C forms with adenosine as a substrate, respectively. The relative substrate specificity ratio showed that the B form was very specific for adenosine.In conclusion, the B form adenosine deaminase was found to be dominant in the frog liver. The Michaelis constant, relative substrate specificity ratio, thin layer electrophoresis and isoelectric focusing distinguished between the adenosine deaminase B and C forms. The technique of ammonium sulfate fractionation gave excellent separation between the B and C forms of adenosine deaminase.Ball State UniversityMuncie, IN 47306