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Selected Publication:

Pekarsky, A; Veiter, L; Rajamanickam, V; Herwig, C; Grunwald-Gruber, C; Altmann, F; Spadiut, O.
(2018): Production of a recombinant peroxidase in different glyco-engineered Pichia pastoris strains: a morphological and physiological comparison
MICROB CELL FACT. 2018; 17: FullText FullText_BOKU

Abstract:
The methylotrophic yeast Pichia pastoris is a common host for the production of recombinant proteins. However, hypermannosylation hinders the use of recombinant proteins from yeast in most biopharmaceutical applications. Glyco-engineered yeast strains produce more homogeneously glycosylated proteins, but can be physiologically impaired and show tendencies for cellular agglomeration, hence are hard to cultivate. Further, comprehensive data regarding growth, physiology and recombinant protein production in the controlled environment of a bioreactor are scarce. A Man(5)GlcNAc(2) glycosylating and a Man(8-10)GlcNAc(2) glycosylating strain showed similar morphological traits during methanol induced shake-flask cultivations to produce the recombinant model protein HRP C1A. Both glyco-engineered strains displayed larger single and budding cells than a wild type strain as well as strong cellular agglomeration. The cores of these agglomerates appeared to be less viable. Despite agglomeration, the Man(5)GlcNAc(2) glycosylating strain showed superior growth, physiology and HRP C1A productivity compared to the Man(8-10)GlcNAc(2) glycosylating strain in shake-flasks and in the bioreactor. Conducting dynamic methanol pulsing revealed that HRP C1A productivity of the Man(5)GlcNAc(2) glycosylating strain is best at a temperature of 30 A degrees C. This study provides the first comprehensive evaluation of growth, physiology and recombinant protein production of a Man(5)GlcNAc(2) glycosylating strain in the controlled environment of a bioreactor. Furthermore, it is evident that cellular agglomeration is likely triggered by a reduced glycan length of cell surface glycans, but does not necessarily lead to lower metabolic activity and recombinant protein production. Man(5)GlcNAc(2) glycosylated HRP C1A production is feasible, yields active protein similar to the wild type strain, but thermal stability of HRP C1A is negatively affected by reduced glycosylation.
Authors BOKU Wien:
Altmann Friedrich
Gr├╝nwald-Gruber Clemens

Find related publications in this database (Keywords)
Pichia pastoris
SuperMan(5)
OCH1
Bioreactor
Cellular agglomeration
Flow cytometry
Glycosylation
Horseradish peroxidase
Morphology


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