Purification of hyperthermophilic archaeal amylolytic enzyme (MJA1) using thermoseparating aqueous two-phase systems.
Purification of a recombinant, thermostable alpha-amylase (MJA1) from the hyperthermophile, Methanococcus jannaschii, was investigated in the ethylene oxide-propylene oxide random copolymer (PEO-PPO)/(NH(4))(2)SO(4), and poly(ethylene glycol) (PEG)/(NH(4))(2)SO(4) aqueous two-phase systems. MJA1 partitioned in the top polymer-rich phase, while the remainder of proteins partitioned in the bottom salt-rich phase. It was found that enzyme recovery of up to 90% with a purification factor of 3.31 was achieved using a single aqueous two-phase extraction step. In addition, the partition behavior of pure amyloglucosidase in polymer/salt aqueous two-phase systems was also evaluated. All of the studied enzymes partitioned unevenly in these polymer/salt systems. This work is the first reported application of thermoseparating polymer aqueous two-phase systems for the purification of extremophile enzymes.
Li
Mian
M
Department of Chemical and Biochemical Engineering, The University of Iowa, 4403 Bowen Science Building, Iowa City, IA 52242-1109, USA. mianl@uwyo.edu
Peeples
Tonya L
TL
D002853:Chromatography, Liquid / Q000379:methods*; D017017:Methanococcus / Q000201:enzymology*; D011994:Recombinant Proteins / Q000302:isolation & purification; D000516:alpha-Amylases / Q000302:isolation & purification*
D011994:Recombinant Proteins; D000516:alpha-Amylases