A simple and nondestructive method for the separation of polysaccharides from beta-glucosidase produced extracellularly by Aspergillus niger.
An apparatus based on electrophoresis has been devised that removes noncovalently bound polysaccharides from extracellular proteins of Aspergillus niger with concomitant partial beta-glucosidase purification and concentration. The apparatus consists of a series of three chambers separated by polyacrylamide gels. Dialyzed and concentrated crude extract of Aspergillus niger containing beta-glucosidase was poured into the middle chamber, while smaller anodic and cathodic chambers contained buffer. When electric current was applied, negatively charged protein-polysaccharide complexes moved toward the anode. Most of the negatively charged proteins, including beta-glucosidase, crossed the gel barrier into the anodic compartment, while neutral polysaccharides were either trapped in the gels or remained in the middle chamber. In this way, 125 ml of dialyzed and concentrated crude extract of Aspergillus niger was processed. Therefore, after 24 h of electrophoresis, 68% of the proteins and 90% of the beta-glucosidase activity, but only negligible amounts of polysaccharide, were transferred to the anodic chamber. The removal of high-molecular-weight polysaccharide from beta-glucosidase had a detrimental effect on the stability of the enzyme.
Siddiqui
K S
KS
National Institute for Biotechnology and Genetic Engineering (NIBGE), Faisalabad, Pakistan.
Rashid
M H
MH
Shemsi
A M
AM
Rajoka
M I
MI
D001234:Aspergillus niger / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel / Q000295:instrumentation / Q000379:methods*; D004795:Enzyme Stability; D005615:Freezing; D011134:Polysaccharides / Q000302:isolation & purification*; D013997:Time Factors; D001617:beta-Glucosidase / Q000096:biosynthesis / Q000737:chemistry / Q000302:isolation & purification*
D011134:Polysaccharides; D001617:beta-Glucosidase