Reddy, Venugopala AR and Ananthanarayanan, VS and Rao, Appaji N (1979) Studies on nucleotidases in plants: Fluorescence and kinetic properties of nucleotide pyrophosphatase from mung bean (Phaseolus aureus) seedlings. In: Archives of Biochemistry and Biophysics, 198 (1). pp. 89-96.
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Nucleotide pyrophosphatase of mung bean seedlings has earlier been isolated in our laboratory in a dimeric form (Mr 65,000) and has been shown to be converted to a tetramer by AMP and to a monomer by p-hydroxymercuribenzoate. All the molecular forms were enzymatically active with different kinetic properties. By a modified procedure using blue-Sepharose affinity chromatography, we have now obtained a dimeric form of the enzyme which is desensitized to AMP interaction. The molecular weight of the desensitized form of the enzyme was found to be the same as that of the native dimeric enzyme. However, the desensitized enzyme functioned with a linear time course, contrary to the biphasic time course exhibited by the native enzyme. In addition, it was not converted to a tetramer on the addition of AMP, had only one binding site for adenine nucleotides, and p-hydroxy-mercuribenzoate had no effect on the time course of the reaction or on the molecular weight of the enzyme. The temperature optimum of the desensitized enzyme was found to be 67 °C in contrast to the optimum of 49 °C for the native dimer. Fifty percent of the tryptophan residues of the desensitized enzyme were not accessible for quenching by iodide. Fluorescence studies gave Kd values of 0.34, 2.2, and 0.8 mImage for AMP, ADP, and ATP, which were close to the Ki values of 0.12, 2.2, and 0.9 mImage , respectively, for these nucleotides. The binding and inhibition studies with AMP and its analogs showed that the 6-amino group and the 5′-phosphate group were essential for the inhibition of the enzyme activity.
|Item Type:||Journal Article|
|Additional Information:||Copyright of this article belongs to Elsevier Science.|
|Department/Centre:||Division of Biological Sciences > Biochemistry
Division of Biological Sciences > Molecular Biophysics Unit
|Date Deposited:||27 Dec 2010 13:05|
|Last Modified:||27 Dec 2010 13:05|
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