HIGH-PERFORMANCE LIQUID-CHROMATOGRAPHY OF BACILLUS-CIRCULANS PEPTIDOGLUTAMINASE FOR LABORATORY AND INDUSTRIAL USES

Gel permeation and anion-exchange chromatography were used to develop methodology for large-scare production of peptidoglutaminase (PGase) from Bacillus circulans cell extract. Sample load, flow-rate and elution profiles were optimized to obtain a highly active DNA-free PGase preparation in high yield. PGase was fractionated into I and II and purified up to 1430-fold for enzymatic determination of glutamine, molecular cloning and protein deamidation research. PGases were also separated directly from B. circulans extract (20-30 mg) in one peak with an 8-fold purification on a 43-ml anion-exchange column at 2 cm/min in 35-40 min. More than 65% of the cell extract proteins were eluted after the PGase peak and contained all the nucleic acids. This method appears to meet the requirements of purity, yield, speed and other economic aspects for successful production of PGase for potential modification of food proteins in industrial reactors.