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Gewählte Publikation:

Pfeffer, M; Maurer, M; Stadlmann, J; Grass, J; Delic, M; Altmann, F; Mattanovich, D; .
(2012): Intracellular interactome of secreted antibody Fab fragment in Pichia pastoris reveals its routes of secretion and degradation.
Appl Microbiol Biotechnol. 2012; 93(6):2503-2512 FullText FullText_BOKU

Abstract:
Protein translation, translocation, folding, processing, and secretion in eukaryotic cells are complex and not always straightforward processes, e.g., different routes of secretion and degradation exist. Formation of malfolded proteins in the endoplasmic reticulum (ER) can be one of the major bottlenecks for recombinant protein production. In this regard, an in-depth analysis of the interactions of a secreted protein during its pathway through the cell may be beneficial, as realized in this study for the methylotrophic yeast Pichia pastoris. The antibody fragment Fab3H6 used here is the anti-idiotype to the HIV neutralizing antibody 2F5 and is known to be intracellularly degraded in significant amounts when expressed in P. pastoris. The interactome of Fab3H6 was analyzed by using a pull-down mass spectrometry approach, and 23 proteins were found to bind specifically to the antibody fragment. Those allowed concluding that Fab3H6 is post-translationally translocated into the ER and degraded via the proteasome as well as the vacuole. In line with this, the expression of Fab3H6 increased the proteasomal activities by over 20%. Partial inhibition of the proteasome resulted in a significant increase of extracellular Fab3H6. Thus, it seems that ER quality control overshoots its requirements for the recombinant protein expressed and that more than just terminally malfolded protein is degraded by ER-associated degradation. This work will further facilitate our understanding how recombinant proteins behave in the secretory pathway.
Autor*innen der BOKU Wien:
Altmann Friedrich
Delic Marizela
Grass Josephine
Mattanovich Diethard
Pfeffer Martin
Stadlmann Johannes
BOKU Gendermonitor:

Find related publications in this database (using NML MeSH Indexing)
Immunoglobulin Fab Fragments/genetics
Immunoglobulin Fab Fragments/metabolism*
Pichia/genetics
Pichia/metabolism*
Protein Binding
Protein Transport
Proteolysis
Recombinant Proteins/genetics
Recombinant Proteins/metabolism
Secretory Pathway*

Find related publications in this database (Keywords)
Interactome
Heterologous protein
Antibody fragment
Secretion
Proteasome
Pichia pastoris


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