Production of a functional human acid maltase in tobacco seeds; biochemical analysis, uptake by human GSDII cells and in vivo studies in GAA knockout mice.

Genetic deficiency of acid alpha glucosidase (GAA) results in glycogen storage\r\ndisease type II (GSDII) or Pompe’s disease. To investigate whether we could generate a\r\nfunctional recombinant human GAA enzyme (tobrhGAA) in tobacco seeds for future\r\nenzyme replacement therapy, we subcloned the human GAA cDNA into the plant expression\r\nplasmid-pBI101 under the control of the soybean β-conglycinin seed-specific promoter and\r\nbiochemically analyzed the tobrhGAA. Tobacco seeds contain the metabolic machinery that\r\nis more compatible with mammalian glycosylation−phosphorylation and processing. We\r\nfound the tobrhGAA to be enzymatically active was readily taken up by GSDII fibroblasts\r\nand in white blood cells from whole blood to reverse the defect. The tobrhGAA corrected the\r\nenzyme defect in tissues at 7 days after a single dose following intraperitoneal (IP) administration\r\nin GAA knockout (GAA−/−) mice. Additionally, we could purify the tobrhGAA since it\r\nbound tightly to the matrix of Sephadex G100 and can be eluted by competition with maltose.