Purification stepAll the purificati

Purification step
All the purification procedures were carried out at
4 ℃ . The pH of culture filtrate (2100 ml) was adjusted
to 5.9 and the filtrate was mixed with an adsorbent
ion-exchanger, DEAE-Sepharose CL-6B (20 ml), in
a beaker. After the crude enzyme was extracted from
DEAE-Sepharose CL-6B with 50 ml of 10 mM acetate
buffer, pH 5.5, containing 0.5 M NaCl, the eluate was
desalted, concentrated, and loaded on a DEAE-Toyopearl
column in FPLC system (Waters 650). The enzyme was
eluted with a linear 0-0.5 M NaCl gradient in 20 mM
acetate buffer, pH 5.5, and fractions with MnP activity
were collected, desalted, and concentrated.