Editing Yeast Fermentation (section)

=== Deglycosylation of recombinant glycoproteins expressed in ''P''. ''pastoris'' using the recombinant Endo H ===

For the post-fermentation treatment, the recombinant ''P''. ''pastoris'' strains producing endo-1, 4-β-mannosidase, phytase and ''Dpn''I were incubated in 100 ml BMGY until the OD<sub>600</sub> value reached approximately 20. All cells were harvested by centrifugation, transferred to 25 ml BMMY and then induced with 1% (v/v) methanol for 144 h. The cell culture supernatant of the ''P''. ''pastoris'' producing recombinant Endo H-P was prepared using the same procedure. The supernatant containing each glycoprotein was mixed with that containing recombinant Endo H-P at a ratio of 10:1 and incubated at 37°C for 1 h.

For co-fermentation, the recombinant ''P''. ''pastoris'' strains expressing DNase I and phytase were separately inoculated in 100 ml BMGY and incubated until the OD<sub>600</sub> value reached approximately 20. All cells were collected from these two samples and transferred into two flasks containing 25 ml BMMY in each. Cells from a 10-ml culture of recombinant Endo H-P-expressing ''P''. ''pastoris'' prepared under the same conditions were collected and added to both flasks, which were then induced with 1% (v/v) methanol for 144 h.
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