Abstract
This is the results of the studies to observe the effect of cultural conditions on the pectin-methyl-esterase (PME) production, and also difference in PME producing activities among various isolates, in Erwinia carotovora.
The organisms were grown in a synthetic pectin medium containing pectin 5g, (NH4)2SO4 13g, KH2PO4 4.5g, Na2HPO4⋅12H2O 2.0g, MgSO4⋅7H2O 0.2g per liter, adjusted to pH 7.0 by NaOH.
The measurement of PME activity was made by the Smith's method, using 4.5ml of pectin substrate and 0.5ml of enzyme solution instead of 7.4ml and 0.6ml, respectively, of Smith's original prescription. 0.5ml aliquot of bacterial culture as the enzyme solution was pipetted into each of two test tubes, and one of them was heated at 100°C for 5min. to inactivate the enzyme. Pectin substrate containing pectin 5g, phenol 2g, NaCl 5.8g per liter was adjusted, right prior to use, to pH7.0 with the addition of NaOH and B. T. B. 4.5ml of this substrate was added to each of the above test tubes, and they were incubated for 24hrs. at 30°C. The pH of the content of these tubes were measured by a glass-electrode pH meter, and the difference in pH value between both tubes was taken as the index of PME activity.
The surface area of the culture contacting with air and also initial pH were found to be the most important among several factors which control the production of PME. Most isolates of E. carotovora could secrete only a little PME in a test tube 1.5cm in diameter, containing 10ml of culture medium.
Despite negligible difference in the growth rate of the bacteria, the larger the surface of the culture contacting with air, the higher was the PME activity evidenced in static culture. An only exception was shown by isolate 551 (E. carotovora f. sp. ananas), which constantly produced abundant enzyme regardless of the surface area of the culture.
In most isolates except 551, this effect of surface area on PME production was increased by low pH of the medium, that is, shift from neutral to acid side considerably lowered the amount of enzyme produced.
Although PME enzyme was most abundantly secreted in the medium when pectin was used as the sole carbon source, pectic acid served fairly effectively as the C-source for the PME enzyme production under cultural conditions having larger surface contacting with air. The low activity of PME was also evidenced in the medium containing glucose as the sole carbon source, but in most isolates except 551 the activity decreased with the lapse of days.
The sort of medium for the pre-culture had little significance as to the amount of PME enzyme produced in the pectin medium.
Under aerobic conditions, all the isolates showing abundant growth in the pectin medium produced PME enzyme, but such isolates as CP or BA needed 2 or 3 days before they showed growth and enzyme production.
The weakly pathogenic isolate P2 showed a very feeble growth and no enzyme production under any cultural conditions.
