The tonoplast H+-pyrophosphatase of radish seedlings: biochemical characteristics |
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Authors: | Maria Chiara Pugliarello Franca Rasi-Caldogno Maria Ida De Michelis Claudio Olivari |
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Institution: | Centro di Studio del C. N. R. suNa Biologia Cellulare e Molecolare delle Piante, Dipartimento di Biologia, Univ. degli Studi di Milano, Via Celoria 26, 1–20133 Milano, Italy;Istituto di Botanica, Univ. di Messina, C.P. 58, I–98166 Messina - S. Agata, Italy. |
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Abstract: | The activity of the H+-pyrophosphatase (H+-PPase) was characterized in microsomes from 24-h-old radish ( Raphanus sativus L., ev. Tondo Rosso Quarantino) seedlings, which are virtually devoid of the tonoplast H+-ATPase. The H+-PPase was localized to membranes which roughly comigrated with the plasma membrane in a sucrose density gradient, but clearly separated from plasma membrane when microsomes were partitioned in an aqueous dextran-polyethylene glycol two-phase system. The H+-PPase activity was strictly dependent on Mg2+ and on the presence of a monovalent cation (K+=Rb+=NH3+Cs+?Na+Li+) and was insensitive to anions such as Cl?, Br?, NO3? and SO42-. It was inhibited by F?, imidodiphosphate and Ca2+. It had a pH optimum between pH 7.5 and 8.5 and was saturated by low concentrations of pyrophosphate (half saturation at 30 μ M pyrophosphate). All of these characteristics are identical to those reported for the tonoplast H+-PPase from various plant materials. The functional molecular weight of the H+-PPase, measured with the radiation-inactivation technique was 96 kDa. |
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Keywords: | H+ pumping H+-pyrophosphatase membrane vesicles radiation-inactivation radish Raphanus sativus L tonoplast |
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