Multiple roles of the cellulase CBHI in enhancing production of fusion antibodies by the filamentous fungus Trichoderma reesei.
The production of Fab antibody fragments in Trichoderma reesei can be increased over 50-fold by fusing the core-linker region of the T. reesei cellulase CBHI (cellobiohydrolase I) to the heavy Fd chain (Nyyssönen et al. 1993). This beneficial role of CBHI in antibody production has now been studied further by comparisons of T. reesei trains producing the light chain only. Fab or CBHI-Fab all of which exhibited identical light chain integration. The N-terminal fusion of CBHI to the heavy Fd chain not only aided secretion, as expected, but also increased the level of mRNA encoding the CBHI-heavy Fd chain, either by stabilizing the messenger or by enhancing transcription. The CBHI part appeared to facilitate secretion at least by aiding the passage through the endoplasmic reticulum, since processing of the signal peptide of the antibody chains seemed to be most efficient in the strain producing CBHI-Fab in contrast to the strains producing light chain or Fab fragment. Interestingly, CBHI core-linker protein, originating from the CBHI-heavy Fd chain, was found in large amounts in the culture medium. The cleavage resulting in this tailless CBHI occurred inside the cell. This suggests that, by omitting the heterologous tail, the secretion of the resulting CBHI core-linker protein is enhanced to a level comparable with secretion of the extracellular T. reesei proteins.
VTT Biotechnology and Food Research, Finland.
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Nyyssönen
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Keränen
D043366:Cellulose 1,4-beta-Cellobiosidase; D003001:Cloning, Molecular; D003470:Culture Media; D005822:Genetic Vectors; D006026:Glycoside Hydrolases / Q000378:metabolism*; D007140:Immunoglobulin Fab Fragments / Q000096:biosynthesis / Q000235:genetics*; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D014242:Trichoderma / Q000235:genetics*
D003470:Culture Media; D007140:Immunoglobulin Fab Fragments; D012333:RNA, Messenger; D011993:Recombinant Fusion Proteins; D006026:Glycoside Hydrolases; D043366:Cellulose 1,4-beta-Cellobiosidase