Purification and properties of polygalacturonic acid trans-eliminase from Bacillus stearothermophilus.

A strain of thermophilic bacteria, Bacillus stearothermophilus, with pectolytic activity has been isolated. It produced an endo-polygalacturonic acid trans-eliminase (endo-PATE, EC 4.2.2.2) extracellularly when grown at 65 degrees C on a pectic acid medium. The PATE was purified 62-fold by the rapid affinity chromatographic method on a Sepharose-polygalacturonamide linked matrix. The absorbed PATE was eluted from the column with a continuous gradient of 0-10(-3)M ethylenediaminetetraacetic acid (EDTA) in phosphate buffer at pH 7.6. The endo-PATE of this organism was much more heat stable than similar enzymes from the mesophilic Bacillus polymyxa and the thermotolerant Bacillus pumilus. The maximum activity of the enzyme occurred at 70 degrees C. With pectic acid as the substrate, the endo-PATE had an optimal pH of 9.0, the highest optimal pH compared with those of similar enzymes from other species of the genus. The molecular weight of the endo-PATE, as determined by chromatography on a Sephadex G-100 gel column, was 24 000.