Balakrishnan, C. V. ; Vaidyanathan, C. S. ; Appaji Rao, N. (1977) Studies on nucleotidases in plants: isolation and properties of the monomeric form of the crystalline and homogeneous mung bean nucleotide pyrophosphatase European Journal of Biochemistry, 78 (1). pp. 95-102. ISSN 0014-2956
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Official URL: http://www3.interscience.wiley.com/journal/1196253...
Related URL: http://dx.doi.org/10.1111/j.1432-1033.1977.00095.pp.x
Abstract
Mung bean nucleotide pyrophosphatase isolated in a crystalline and homogeneous form as a dimer with a molecular weight of 65000 was converted by AMP into a tetramer. The tetramer was enzymatically active with altered kinetic properties. This conversion of the dimeric form by AMP to a tetrameric one was prevented by treating the dimer with p-hydroxymercuribenzoate. The molecular weight of the p-hydroxymercuribenzoate-treated enzyme was determined to be 32700 by a combination of Stokes' radius (2.4 nm) and sedimentation velocity (S20,w = 1.9 S), by thin-layer gel chromatography on superfine Sephadex G-200 and by sodium dodecylsulfate/polyacrylamide gel electrophoresis. The monomer obtained by treatment of the native enzyme with p-hydroxymercuribenzoate was isolated by passage of the dissociated enzyme through a column of Biogel P-200. The monomer was optimally active at 37°C, whereas the dimer and tetramer were active at 49°C. All the three enzyme forms were maximally active at pH 9.4. The Km and V (measured as rate of FAD hydrolysis per mg protein) for FAD of the three enzyme forms were for the monomer, 0.5mM and 7.0 μmolmin-1, for the dimer, 0.25mM and 3.3 μmolmin-1 and for the tetramer, 0.58mM and 2.5 μmolmin-1, respectively. The time course of the reaction of the monomer was linear and comparable to the initial fast rate of the dimer. The monomer was not converted to a tetramer or a dimer on the addition of AMP; and it was irreversibly inhibited by urea and EDTA. ATP and ADP were noncompetitive inhibitors of the monomer.
Item Type: | Article |
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Source: | Copyright of this article belongs to John Wiley and Sons, Inc. |
ID Code: | 21265 |
Deposited On: | 20 Nov 2010 13:12 |
Last Modified: | 17 May 2016 05:28 |
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