Aqueous extraction of beta-glucuronidase from transgenic canola: kinetics and microstructure.

Abstract

Aqueous extraction kinetics of recombinant beta-glucuronidase (rGUS) from transgenic canola (Brassica napus) was investigated in terms of the particle size and microstructural characteristics resulting from canola seed processing. The canola had been transformed to express recombinant GUS intracellulary in the seed, and electron microscopy showed that the cells are distributed among (1) disrupted cells in a thin layer at or adjacent to the particle surface, (2) disrupted cells within the interior, and (3) intact cells within the interior. A simple compartmental model containing two extractable pools and a third nonextractable pool fitted the batch extraction results very well. Comparing the rate constants from the model to estimates of expected transport rates from the observed cell fractions showed that the two extractable pools roughly correspond to the two disrupted cell fractions. Both flaking, causing more extensive cell wall damage throughout the seed, and grinding, increasing the total surface area, increase the size of the first pool and, therefore, the extraction yield. Mass transfer from the same type of pool from two types of processed seed behaved similarly. GUS extraction from the first extractable pool is 10-20 times faster (<1 min) than from the second extractable pool.