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High-efficiency secretory production of peroxidase C1a using vesicular transport engineering in transgenic tobacco

Paper ID Volume ID Publish Year Pages File Format Full-Text
22139 43256 2006 8 PDF Available
Title
High-efficiency secretory production of peroxidase C1a using vesicular transport engineering in transgenic tobacco
Abstract

Horseradish peroxidase isozyme C1a (HRP C1a) is widely used as a reporter enzyme in a variety of detection procedures such as enzyme-linked immunosorbent assay (ELISA) and western blotting. We previously isolated the gene encoding HRP C1a and showed that HRP C1a is at first translated as a preproprotein containing propeptides at its N- and C-termini (N-terminal secretion signal peptide and C-terminal propeptide; CTPP). The signal peptide (sp) is necessary for endoplasmic reticulum (ER) translocation and the CTPP acts as a vacuolar sorting determinant. Furthermore, HRP C1a was secreted into the culture medium from cells expressing the HRP C1a gene without the CTPP region. We optimized the secretory production system of HRP C1a in tobacco plants. To determine a suitable signal peptide for high-efficient secretion of proteins, three types of sp derived from HRP C1a (C1Psp), β-d-glucan exohydrolase (GEsp) and 38 kDa peroxidase (38Psp) were compared. GE and 38P are secretory proteins highly accumulated in the culture medium of BY2 cells. The secretion efficiency was increased by 34% and 53% when GEsp and 38Psp was used, respectively. Next, we used a translational enhancer, the 5′-untranslated region of Nicotiana tabacum alcohol dehydrogenase gene (NtADH 5′-UTR). The production of HRP C1a was increased by placing NtADH 5′UTR in front of the ORF in BY2 cells. These results indicate that the localization and expression level of recombinant proteins can be controlled by the use of propeptides and 5′UTR, respectively. Finally, high-efficiency secretory production of the HRP C1a was also achieved in transgenic tobacco.

Keywords
horseradish peroxidase; signal peptides; translational enhancer; Nicotiana tabacum; cultured tobacco cell
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High-efficiency secretory production of peroxidase C1a using vesicular transport engineering in transgenic tobacco
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Publisher
Database: Elsevier - ScienceDirect
Journal: Journal of Bioscience and Bioengineering - Volume 102, Issue 2, August 2006, Pages 102–109
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