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Coordinate expression of multiple proteins in plant cells by exploiting endogenous kex2p-like protease activity.
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- Author(s): Zhang B;Zhang B; Rapolu M; Huang L; Su WW
- Source:
Plant biotechnology journal [Plant Biotechnol J] 2011 Dec; Vol. 9 (9), pp. 970-81. Date of Electronic Publication: 2011 Mar 28.
- Publication Type:
Journal Article; Research Support, U.S. Gov't, Non-P.H.S.
- Language:
English
- Additional Information
- Source:
Publisher: Wiley on behalf of the Society for Experimental Biology, Association of Applied Biologists Country of Publication: England NLM ID: 101201889 Publication Model: Print-Electronic Cited Medium: Internet ISSN: 1467-7652 (Electronic) Linking ISSN: 14677644 NLM ISO Abbreviation: Plant Biotechnol J Subsets: MEDLINE
- Publication Information:
Publication: 2014- : Oxford Wiley on behalf of the Society for Experimental Biology, Association of Applied Biologists
Original Publication: [Oxford] : Blackwell Pub., c2003-
- Subject Terms:
- Abstract:
Simultaneous expression of multiple proteins in plants finds ample applications. Here, we examined the biotechnological application of native kex2p-like protease activity in plants for coordinate expression of multiple secretory proteins from a single transgene encoding a cleavable polyprotein precursor. We expressed a secretory red fluorescent protein (DsRed) or human cytokine (GMCSF), fused to a downstream green fluorescent protein (GFP) by a linker containing putative recognition sites of the kex2p-like protease in tobacco cells and referred to them as RKG and GKG cells, respectively. Our analyses showed that GFP is cleaved off the fusion proteins and secreted into the media by both RKG and GKG cells. The cleaved GFP product displayed the expected fluorescence characteristics. Using GFP immunoprecipitation and fluorescence analysis, the cleaved DsRed product in the RKG cells was found to be functional as well. However, DsRed was not detected in the RKG culture medium, possibly due to its tetramer formation. Cleaved and biologically active GMCSF could also be detected in GKG cell extracts, but secreted GMCSF was found to be only at a low level, likely because of instability of GMCSF protein in the medium. Processing of polyprotein precursors was observed to be similarly effective in tobacco leaf, stem and root tissues. Importantly, we also demonstrated that, via agroinfiltration, polyprotein precursors can be efficiently processed in plant species other than tobacco. Collectively, our results demonstrate the utility of native kex2p-like protease activity for the expression of multiple secretory proteins in plant cells using cleavable polyprotein precursors containing kex2p linker(s).
(© 2011 The Authors. Plant Biotechnology Journal © 2011 Society for Experimental Biology, Association of Applied Biologists and Blackwell Publishing Ltd.)
- Accession Number:
0 (Culture Media)
0 (Enzyme Precursors)
0 (Luminescent Proteins)
0 (Polyproteins)
0 (Protein Sorting Signals)
0 (Recombinant Fusion Proteins)
0 (Saccharomyces cerevisiae Proteins)
147336-22-9 (Green Fluorescent Proteins)
83869-56-1 (Granulocyte-Macrophage Colony-Stimulating Factor)
EC 3.4.21.- (Proprotein Convertases)
EC 3.4.21.61 (KEX2 protein, S cerevisiae)
- Publication Date:
Date Created: 20110330 Date Completed: 20120227 Latest Revision: 20231213
- Publication Date:
20231215
- Accession Number:
10.1111/j.1467-7652.2011.00607.x
- Accession Number:
21443545
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