Extracellular and Intracellular Exopectate Lyases of Erwinia carotovora subsp. carotovora

A strain of Erwinia carotovora was found to produce an exopectate lyase (exo-PAL) both in the culture fluid and in the cells. Each activity of the extracellular and intracellular exo-PALs fell into two fractions by cellulose ion exchanger chromatographies. They were named tentatively enzyme 1-A and 1-B, and 2-A and 2-B, for the former and the latter, respectively. As the amount of enzyme 1-A was very small, it was discarded. The yields of enzyme 1-B, 2-A, and 2-B were 21.6, 21.4, and 14.0 %, respectively; the purification degree of each enzyme was considerably high. These three exo-PALs resembled one another very closely in the following respects: (1) The optimum pH was about 9 in both Tris-HC1 and borate buffers. (2) Na+ was an effective activator; its optimum concentration was about 15 mM. Mn2+ and Co2+ stimulated weakly regardless of the presence or absence of Na+, but Ca2+ showed a weak stimulation only in the absence of Na+. (3) The exo-PALs were inhibited to varying extent by Cu2+, Hg2+, Sr2+, Mg2+, and Ba2+. (4) Addition of 1 mM EDTA led to a total loss of activity. From these results, we considered that both the extracellular and intracellular exo-PALs are probably identical. As enzyme source, the intracellular exo-PAL is superior to the extracellular exo-PAL because of the high recovery and ease of preparation of the former enzyme.