Enhancement of α-galactosidase production using novel Actinoplanes utahensis B1 strain: sequential optimization and purification of enzyme

Abstract

α-Galactosidase is an important exoglycosidase belonging to the hydrolase class of enzyme, which has therapeutic and industrial potential. It plays a crucial role in hydrolyzing α-1,6 linked terminal galacto-oligosaccharide residues such as melibiose, ra nose, and branched polysaccharides such as Galacto-glucomannans and Galactomannans. In this study Actinoplanes utahensis was explored for α-Galactosidase production, yield improvement and activity enhancement by puri cation. Initially nine media components were screened by using the Plackett-Burman design (PBD). Among these components, sucrose, soya bean our, and sodium glutamate were identi ed as best supporting nutrients for highest enzyme secretion by A. utahensis. Later, the Central Composite Design (CCD) was implemented to ne-tune the optimization of these components. Based on sequential statistical optimization methodologies, a signi cant, 3.64-fold increase in α-galactosidase production, from 16 to 58.37 U/mL was achieved.The enzyme was puri ed by ultra ltration-I followed by multimode chromatography and ultra ltration-II. The purity of the enzyme was con rmed by Sodium Dodecyl Sulphate -Polyacrylamide Agarose Gel Electrophoresis (SDS-PAGE) which revealed a single distinctive band with a molecular weight of approximately 72kDa. Additionally, it was determined that this process resulted in a 2.03fold increase in purity. The puri ed α-galactosidase showed an activity of 2304 U/mL with a speci c activity of 288 U/mg. This study demonstrates the feasibility of scalable α-galactosidase production, which has various industrial applications.