Separation of Thermostable Pectinmethylesterase from Marsh Grapefruit Pulp

A rapid, ion-exchange method that isolates thermostable pectinmethylesterase (TS-PME) from Marsh grapefruit pulp is presented. TS-PME was selectively extracted with 1 M NaCl and equilibrated at low pH (3.1 +/- 0.04). After dilution to a final concentration of 50 mM acetate buffer, pH 5.5, and 0.1, 0.25, or 0.5 M NaCl, the extract was applied to an ion-exchange column without ammonium sulfate precipitation and dialysis. The percent yield varied from an average of 70% (0.1 M NaCl) to 14% (0.5 M NaCl), with a specific activity of >400 U/mg protein in the latter sample. TS-PME activity of some column fractions varied compared with crude extracts before column separation. Some fractions lost thermostability after separation if loaded at 0.1 M or 0.25 M NaCl. In extracts that were loaded at 0.5 M NaCl, the TS-PME activity was significantly higher than that observed in the heated crude extracts. Ion-exchange chromatography may have separated an unidentified protective factor in PME fractions.