| The genetic disorder mucopolysaccharidosis IIIB, which primarily affects the central nervous system (CNS), is caused by a deficiency in the enzyme alpha-N-acetylglucosaminidase (Naglu). Recombinant Naglu is unable to enter cells or cross the blood-brain barrier (BBB), making potential enzyme replacement therapy infeasible. To enable Naglu to be endocytosed by cells and perhaps cross the BBB, two fusion proteins of Naglu with the HIV-1 Tat protein transduction domain (PTD) or a modified PTD were created. This project explored the use of a Spodoptera frugiperda 9 ( Sf9) expression system utilizing the p2ZoptcxF vector to produce and purify active Naglu and active Naglu-PTD fusion proteins. It was found that the Sf9 expression system produced active Naglu, that the addition of the PTD fusion moieties did not decrease its activity, and that it was possible to purify this protein to near homogeneity. |
