ASPARAGINE CATABOLISM IN BRYOPHYTES - PURIFICATION AND CHARACTERIZATION OF 2 L-ASPARAGINASE ISOFORMS FROM SPHAGNUM-FALLAX

Nitrogen represents a critical nutrient in raised bogs. In Sphagna, do minating this habitat, the prevalent storage amino acid asparagine is catabolized predominantly by the enzyme L-asparaginase (EC 3.5.1.1), L -asparaginase activity has been detected in each of 10 Sphagnum specie s investigated. In Sphagnum fallax Klinggr. (Klinggr, clone 1) cultiva ted under axenic conditions in continuous feed bioreactors, the enzyme displayed a light dependent increase in activity. We separated two is oforms, designated L-asparaginase 1 and 2, characterized by their diff erent elution patterns from an enionexchange column. In stem segments only L-asparaginase 2 could be detected, whereas in capitulae L-aspara ginase 1 represented the dominating isoform. Purified chloroplasts dis played no L-asparaginase activity. Almost the entire activity was loca ted in the cytosolic fraction. L-asparaginase 1 and 2 have been purifi ed 82-fold and 188-fold, respectively, by ion-exchange, size-exclusion and hydrophobic interaction chromatography. Identical pH optima (8.2) and molecular weights (126 000) were determined. The K-m values for a sparagine (7.4 mM 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-asparaginases of microorganisms. So, the characteristics of the bry ophyte enzyme appear to be intermediate between those from higher plan ts and those from microorganisms.