Utilization of Kifunensine and Sorbitol for Enhanced Production of Glyco-engineered Acid α-Glucosidase in Transgenic Rice Cell Cultures

Production of recombinant acid -glucosidase (GAA) with high-mannose glycan and least plant-specific residues in transgenic rice cell cultures is necessary for effective treatment of Pompe disease, a lysosomal storage disorder, using enzyme replacement therapy.Kifunensine was used as a regulator of N-glycosylation with plant expression system and it showed negative effect on the expression level of GAA.In order to increase protein expression with the presence of kifunensine, sorbitol was used as an alternative carbon source under sugar-depleted environment to enhance both cell viability and GAA production.Concentrations of kifunensine and sorbitol were optimized and the combined use of 2.5 mM kifunensine and 40 mM sorbitol was found to be the best for cell growth, viability and GAA expression.The content of high-mannose type glycan was 3.8% in control culture and 62% in 2.5 mM kifunensine alone.It could be improved up to 77.9% by the combined supplementation of 40 mM sorbitol.Therefore, it was clear that the combined use of kifunensine and sorbitol can improve GAA expression as well as the content of high-mannose type glycan.