Summary: Extracellular lipase was purified from a Tween 80-limited continuous culture of EF2 by ultrafiltration of the culture supernatant followed by anion-exchange and gel-filtration FPLC. The lipase was composed of a single subunit ( 29000, pI 4·9), which was capable of a variable degree of aggregation, and which exhibited both lipase activity, measured with the insoluble substrate olive oil (predominantly triolein), and esterase activity, measured with the soluble substrates -nitrophenyl acetate and Tween 80. Lipase activity was approximately eight times higher than either type of esterase activity ( , approximately 3000 sfor the hydrolysis of olive oil). The enzyme showed a marked regiospecificity for the 1,3-oleyl residues of radiolabelled triolein, was relatively stable at moderate temperatures (exhibiting a biphasic loss of activity with an initial of 17·5 min at 60 °C) and was very stable to freezing and thawing. Lipase activity was only weakly inhibited by the serine-active reagent 3,4-dichloroisocoumarin, and was not inhibited by the chelating agent EDTA (1 mm). The N-terminal amino acid sequence of the EF2 lipase showed a marked similarity to those of several other bacterial lipases.


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