Table I.
Extract Origin | Yield
|
Purification
Factor
|
Total Activitya
|
|||
---|---|---|---|---|---|---|
sta8 | + | sta8 | + | sta8 | + | |
% | ||||||
Crude extract | 100 | 100 | 0 | 0 | 225 | 322 |
Protamine sulfate | 74 | 65 | 1, 7 | 1, 3 | 167 | 210 |
Ammonium sulfate | 46 | 40 | 2, 4 | 1, 6 | 105 | 130 |
Gel permeation (52–65) | – | 17 | – | 9, 9 | – | 55 |
Gel permeation (58–66) | 19 | – | 9, 9 | – | 42 | – |
Anion exchange (19–20) | – | 2, 5 | – | 140 | – | 8, 2 |
Anion exchange (15–16) | 4, 5 | – | 77 | – | 10, 2 | – |
Purification was from 4 × 1010 cells. Debranching activity corresponds to micromoles of maltotriose equivalents produced per hour from amylopectin for the whole fraction in study. After the gel permeation step, wild-type isoamylase was free of interfering starch hydrolytic activity, whereas trace amounts of α-amylase can be detected by zymograms in the sta8 semipurified fractions.