The activity of novel uncharacterized extremophilic L-asparaginases from the psychrophilic fungi _Sclerotinia_ _borealis_, the thermoacidophilic crenarchea _Acidilobus saccharovorans,_ and the thermophilic bacteria _Melioribacter roseus_ were studied. Active enzymes were produced via the expression of the native L-asparaginase gene from _M. roseus_ (MrA) and synthetic genes encoding fungal _S._ _borealis_ (SbA) and archeal _A. saccharovorans_ (AsA) L-asparaginases after codon optimization in _Escherichia coli_ cells. In the study, the maximum specific activity at different temperatures and pH was observed for MrA. The activity of MrA crude extract was highest at 75°С and a pH of 9.0. Metal ions (1 mM) differed in their effects on enzyme activity. Сu2+ and Zn2+ ions completely abolished enzyme activity. Changes in the specific activity of MrA crude extract in the presence of Fe3+, Ni2+, Ca2+, and Mg2+ varied within 5-28%. Our findings show that L-asparaginase of _M. roseus_ may be a promising object for the further study of enzymatic properties and biotechnological applications.