Purification and Characterization of Extracellular Alginate Lyase from Enterobacter cloacae M-1

Abstract

An alginate lyase from the culture supernatant of Enterobacter cloacae M-1 was purified by ammonium sulfate precipitation, cation-exchange chromatography (SP-Toyopearl), and gel filtration (Ultrogel AcA44). The final preparation thus obtained showed a single band on SDS-PAGE. The purified enzyme had the molecular weight of 38, 000 and 32, 000 by SDS-PAGE and gel filtration, respectively. The pI of the enzyme was 8. 9. The optimum pH and temperature for the enzyme reaction were around 7. 8 and 30°C, respectively. The enzyme was unstable on heating. EDTA completely inhibited the enzyme activity, but the activity was completely restored by the treatment with CaCl₂. The enzyme was specific for poly-guluronate and produced several kinds of unsaturated oligomers from the gluluronate. This suggested that the enzyme could be classified as an endo poly-guluronate lyase.