Phosphorus metabolism in plants is controlled by numerous acid phosphatases. This diversity makes it difficult to isolate and characterize a single native enzyme, especially that of not dominating form in the testing tissue, from among the others displaying similar activity. Introducing a relatively simple procedure comprising a preparative native PAGE as the main purification step, together with DEAE-Sephacel anion exchange chromatography and gel filtration on Superdex 75, let us to separate two low molecular weight (35 kDa and 42 kDa) acid phosphatases from lupin cotyledons. The first of them was the subject of more detailed study.

The 35 kDa enzyme is a thermostable tartrate-resistant metallophosphatase. It exhibits an affinity for phosphate monoesters of various aryl substrates and for inorganic pyrophosphate in a broad pH range with optimum at pH 5.0. It displays neither phosphodiesterase nor acidic or alkaline phytase activity. The high level of hydrolysis of phosphotyrosine but not phosphoserine, may suggest its possible function in vivo. This form of acid phosphatase was detected in all vegetative organs of the lupin seedlings but not in the dry seed extract. The level of its activity increases in phosphorus-deficient conditions, especially in roots.

Despite the 35 kDa lupin enzyme appears to be an acidic, ConA non-reactive protein, which differentiates it from the animal LMW TRAP (basic glycoproteins), its overall biochemical characteristics is close to this group of enzymes. The potential mammalian-like phosphatase sequences were detected in a wide range of plants [1-4]. Only one protein representing this group, possessing an acid phosphatase activity, has been isolated from phosphate deprived A. thaliana seedlings [4], but its substrate specificity is not known. The conservation of amino acid residues involved in coordination of metal ions in the active centre, as well as similarity of the predicted three-dimensional structure of the plant enzymes to the known structures of the mammalian ones [1] let to presume similar enzymatic activity of both groups. The elucidation of the possible relationship of the isolated lupin enzyme to the plant mammalian-like LMW acid phosphatases is the subject of our ungoing studies.

Reference

1. Schenk G., Guddat L. W., Ge Y., Carrington L. E., Hume D. A., Hamilton S., de Jersey J. (2000). Gene 250, 117-125.
2. Li D., Zhu H., Liu K., Liu X., Leggewie G., Udvardis M., Wang T. (2002) J. Biol. Chem. 277, 27772-27781.
3. Zimmermann P., Regierer B., Kossmann J., Frossard E., Amrhein N., Bucher M. (2004) Plant. Biol. 6, 519-528.
4. del Pozo J. C., Allona I., Rubio V., Leyva A., de la Pena A., Aragoncillo C., Paz-Ares J. (1999) Plant J. 19, 579-589.

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