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Applying unconventional secretion of the endochitinase Cts1 to export heterologous proteins in Ustilago maydis

Paper ID Volume ID Publish Year Pages File Format Full-Text
23789 43473 2012 12 PDF Available
Title
Applying unconventional secretion of the endochitinase Cts1 to export heterologous proteins in Ustilago maydis
Abstract

The demand on the biotechnological production of proteins for pharmaceutical, medical and industrial applications is steadily growing. For the production of challenging proteins, we aim to establish a novel expression platform in the well characterized eukaryotic microorganism Ustilago maydis. In filaments of this fungus, secretion of the endochitinase Cts1 depends on mRNA transport along microtubules, which is mediated by the key RNA-binding protein Rrm4. Here, we report two important findings: (i) Cts1 secretion occurs via a novel unconventional route and (ii) this secretory mechanism can be exploited for the export of active heterologous proteins. Initially, we used β-glucuronidase (Gus) as a reporter for unconventional secretion. This bacterial enzyme is inactivated by N-glycosylation during its passage through the conventional eukaryotic secretory pathway. By contrast, in our system Gus was exported in its active form by fusion to Cts1 confirming its secretion by an unconventional route. As a proof-of-principle for economically important biopharmaceuticals we expressed an active single-chain antibody. Importantly, the novel protein export pathway circumvents N-glycosylation which is advantageous in many applications, e.g., to avoid undesired immune reactions in humans. Thus, the unconventional Cts1 secretion machinery has a high potential for the production of biotechnologically relevant proteins.

► The endochitinase Cts1 of the eukaryotic model organism Ustilago maydis is secreted by an unconventional mechanism. ► The unconventional secretory pathway circumvents N-glycosylation. ► Unconventional secretion of Cts1 can be exploited to co-export heterologous proteins to the culture supernatant.

Keywords
SMART, Simple Modular Architecture Research Tool; TMHMM, transmembrane hidden Markov model; Gus, β-glucuronidase; CM, complete medium; NM, nitrate minimal medium; Gfp, green fluorescent protein; ER, endoplasmic reticulum; scFv, single chain fragment varia
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Applying unconventional secretion of the endochitinase Cts1 to export heterologous proteins in Ustilago maydis
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Publisher
Database: Elsevier - ScienceDirect
Journal: Journal of Biotechnology - Volume 161, Issue 2, 15 October 2012, Pages 80–91
Authors
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Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering
Get Full-Text Now
Don't Miss Today's Special Offer
Price was $35.95
You save - $31
Price after discount Only $4.95
100% Money Back Guarantee
Full-text PDF Download
Online Support
Any Questions? feel free to contact us