Topic
Pichia pastoris
About: Pichia pastoris is a research topic. Over the lifetime, 7937 publications have been published within this topic receiving 162645 citations. The topic is also known as: Komagataella pastoris.
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TL;DR: The GAP promoter based system obviates the need to use and extensively monitor methanol during recombinant antigen production and has the potential for continuous culture wherein several batches of recombinant protein-containing biomass can be harvested from a single initial fermentation.
157 citations
TL;DR: Despite the fact that gelatins are highly susceptible to proteolysis because of their unfolded structure, the recombinant protein was shown to be fully intact by SDS-PAGE, N-terminal sequencing, gel filtration chromatography and mass spectrometry.
Abstract: A custom-designed, highly hydrophilic gelatin was produced in Pichia pastoris. Secreted production levels in single-copy transformants were in the range 3-6 g/l of clarified broth and purification to near homogeneity could be accomplished by differential ammonium sulfate precipitation. Despite the fact that gelatins are highly susceptible to proteolysis because of their unfolded structure, the recombinant protein was shown to be fully intact by SDS-PAGE, N-terminal sequencing, gel filtration chromatography and mass spectrometry. Owing to its highly hydrophilic nature, the migration of the synthetic gelatin in SDS-PAGE was severely delayed. Esterification of the carboxylic amino acid side chains resulted in normal migration. The high polarity of the synthetic gelatin also accounts for its negligible surface activity in water at concentrations up to 5% (w/v), as determined by tensiometry. Circular dichroism spectrometry showed that the non-hydroxylated gelatin did not form triple helices at 4 degrees C. The spectrum was even more representative of the random coil conformation than the spectrum of natural non-hydroxylated gelatins.
156 citations
TL;DR: Both crosslinked diabodies were more stable to heat and the crosslinked bivalent diabody showed improved localization to CEA+ human tumour xenografts in nude mice.
Abstract: Diabodies are dimeric antibody fragments held together by associated heavy and light chain variable domains present on different polypeptide chains. To improve their stability we have introduced cysteine residues into the V-domains to promote the disulphide crosslinking of the dimer. A crosslinked bivalent diabody against carcinoembryonic antigen (CEA) and a crosslinked bispecific diabody against CEA and the T-cell co-receptor CD3 were expressed from Pichia pastoris and Escherichia coli by secretion. From Pichia (but not E.coli) the chains were almost quantitatively crosslinked. Compared with the parent diabodies both crosslinked diabodies were more stable to heat (by >7 degrees C) and the crosslinked bivalent diabody showed improved localization to CEA+ human tumour xenografts in nude mice.
154 citations
TL;DR: Proof of concept that glycoengineered yeast can be used to produce functional full-length monoclonal antibodies at commercially viable productivities is provided.
153 citations
TL;DR: The impressive methodological developments of the last years to model and analyze the metabolism of P. pastoris, and to engineer its genome and metabolic pathways are summarized.
153 citations