H. 1996. Asparagine catabolism in bryophyles: Purification and characterization of two L-asparaginase isoforms from Sphagnum fallax. -Physiol. Plant. 97: 402-410.Nitrogen represents a critical nutrient in raised bogs. In Sptiagna, dominating this habitat, the prevalent storage amino acid asparagine is cataholized predominantly by the enzyme L-asparaginase (EC 3.5.1.1). L-asparaginase activity has been detected in each of 10 Sphagnum species investigated. In Sphagnum fallax Klinggr. (Klinggr. clone 1) cultivated under axenic conditions in continuous feed bioreaclors, the enzyme displayed a light dependent increase in activity. We separated two isoforms, designated L-asparaginase 1 and 2, eharacterized by their different elution patterns from an anionexchange column. In stem segments only L-asparaginase 2 could be detected, whereas in capitulae L-asparaginase 1 represented the dominating isoform. Purified chloroplasts displayed no L-aspai'aginase activity. Almost the entire activity was located in the cytosolic fraction. L-asparaginase 1 and 2 have been purified 82-fold and 188-foId, respectively, by ion-exchange, size-exclusion and hydrophobic interaction chromatography. Identical pH optima (8.2) and molecular weights (126 000) were determined. The Km values for aspaiagine (7.4 \\\M for L-asparaginase 1 and 6.2 mM for L-asparaginase 2) were in the range of those described for higher plants. On the other hand Sphagnum L-asparaginase is comprised of four subunits as are the L-asparagina.ses ot microorganisms. So, the characteristics of the bryophyte enzjine appear to be intermediate between those from higher plants and those from microorganisms.
H. 1997. Ammonium assimilation in bryophytes. L-glutamine synthetase from Sphagnum fallax. -Physiol. Plant. 101: 86-92.Cytosolie and piastidic L-glutamine synthetase (EC 6.3.1.2) isoenzymes from Sphagnum fallax Klinggr. (Klinggr. clone 1) were separated by size-exclusion and ion exchange chromatography. The cytosolie enzyme (GSi) was purified to apparent electrophoretic homogeneity. The native enzyme had a molecular mass of 390 ± 20 kDa as estimated by gel filtration and was apparently composed of 8 subunits with molecular masses of 48 kDa. GS, activity could be measured from pH 6.8 to 8.6 in 50 mM imidazole buffer, with a broad optimum between pH 7.2 and 8.0. The K^ values were 2.5 mM, 0.5 mM and 0.5 mM for L-glutamate, ammonium and ATP, respectively. The enzyme was inhibited by more than 10 mM ammonium or glutamate. The incorporation of "NH4 into amino acids was observed in vivo using '% NMR. Label from ammonium was first detected in the amide N of glutamine, and only subsequently in the amino N of glutamate. Moreover, no assimilation was detected in the presence of the specific GS inhibitor methionine sulfoximine. These observations are consistent with a dominant role for GS in the assimilation of ammonium in Sphagnum.
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