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Research article
Elastin-like polypeptide and γ-zein fusions significantly increase recombinant protein accumulation in soybean seeds
Xiangdong Yang
Jilin Academy of Agricultural Sciences
Corresponding Author
ORCiD: https://orcid.org/0000-0002-7101-1626
License:
This work is licensed under a CC BY 4.0 License. Read Full License
Background
Soybean seeds are an ideal host for the production of recombinant proteins because of their high content of proteins, long-term stability of seed proteins under ambient conditions, and easy establishment of efficient purification protocols.
Results
In this study, a polypeptide fusion strategy was applied to explore the capacity of elastin-like polypeptide (ELP) and γ-zein fusions in increasing the accumulation of the recombinant protein in soybean seeds. Transgenic soybean plants were generated to express the γ-zein- or ELP-fused green fluorescent protein (GFP) under the control of the soybean seed-specific promoter BCSP. Significant differences were observed in the accumulation of zein-GFP and GFP-ELP from that of the unfused GFP in transgenic soybean seeds based on the total soluble protein (TSP), despite the low-copy of T-DNA insertions and similar expression at the mRNA levels in selected transgenic lines. The average levels of zein-GFP and GFP-ELP accumulated in immature seeds of these transgenic lines were 0.99% and 0.29% TSP, respectively, compared with 0.07% TSP of the unfused GFP. In mature soybean seeds, the accumulation of zein-GFP and GFP-ELP proteins was 1.8% and 0.84% TSP, an increase of 3.91- and 1.82-fold, respectively, in comparison with that of the unfused GFP (0.46% TSP). Confocal laser scanning analysis showed that both zein-GFP and GFP-ELP were deposited in the protein storage vacuoles of transgenic seeds, while the unfused GFP was mainly distributed in the cytoplasm. Despite increased recombinant protein accumulation, there were no significant changes in the total protein and oil content in seeds between the transgenic and non-transformed plants, suggesting the possible presence of threshold limits of total protein accumulation in transgenic soybean seeds.
Conclusions
Overall, our results indicate that γ-zein and ELP fusions significantly increased the accumulation of the recombinant protein, but exhibited no significant influence on the total protein and oil content in soybean seeds.
Keywords
Soybean seed, Fusion partner, Elastin-like polypeptide, γ-Zein, Recombinant protein
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CURRENT STATUS: Under Revision
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Posted 30 Dec, 2019
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Editorial decision: Major revision
On 02 Apr, 2020
Review #2 received
Received 14 Feb, 2020
Review #3 received
Received 11 Feb, 2020
Review #1 received
Received 29 Jan, 2020
Reviewer #3 agreed
On 27 Jan, 2020
Reviewer #1 agreed
On 21 Jan, 2020
Reviewer #2 agreed
On 21 Jan, 2020
Reviewers invited
Invitations sent on 20 Jan, 2020
Editor assigned
On 17 Dec, 2019
First submitted
On 16 Dec, 2019
Submission checks complete
On 16 Dec, 2019
Editor invited
On 16 Dec, 2019
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Subject Areas
Biotechnology and Bioengineering
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This preprint is under consideration at BMC Biotechnology. A preprint is a preliminary version of a manuscript that has not completed peer review at a journal. Research Square does not conduct peer review prior to posting preprints. The posting of a preprint on this server should not be interpreted as an endorsement of its validity or suitability for dissemination as established information or for guiding clinical practice.
Learn more about In Review
Research article
Elastin-like polypeptide and γ-zein fusions significantly increase recombinant protein accumulation in soybean seeds
Xiangdong Yang
Jilin Academy of Agricultural Sciences
Corresponding Author
ORCiD: https://orcid.org/0000-0002-7101-1626
Badges
Prescreen
This manuscript passed our Prescreen assessment
Complete author information
Appropriate declaration statements
Potential risks to human health
Prescreen
Peer Review Timeline
CURRENT STATUS: Under Revision
Version 1
Posted 30 Dec, 2019
No community comments so far
Editorial decision: Major revision
On 02 Apr, 2020
Review #2 received
Received 14 Feb, 2020
Review #3 received
Received 11 Feb, 2020
Review #1 received
Received 29 Jan, 2020
Reviewer #3 agreed
On 27 Jan, 2020
Reviewer #1 agreed
On 21 Jan, 2020
Reviewer #2 agreed
On 21 Jan, 2020
Reviewers invited
Invitations sent on 20 Jan, 2020
Editor assigned
On 17 Dec, 2019
First submitted
On 16 Dec, 2019
Submission checks complete
On 16 Dec, 2019
Editor invited
On 16 Dec, 2019
Metrics
Comments: 0
PDF Downloads: ...
HTML Views: ...
Subject Areas
Biotechnology and Bioengineering
License:
This work is licensed under a CC BY 4.0 License. Read Full License
Background
Soybean seeds are an ideal host for the production of recombinant proteins because of their high content of proteins, long-term stability of seed proteins under ambient conditions, and easy establishment of efficient purification protocols.
Results
In this study, a polypeptide fusion strategy was applied to explore the capacity of elastin-like polypeptide (ELP) and γ-zein fusions in increasing the accumulation of the recombinant protein in soybean seeds. Transgenic soybean plants were generated to express the γ-zein- or ELP-fused green fluorescent protein (GFP) under the control of the soybean seed-specific promoter BCSP. Significant differences were observed in the accumulation of zein-GFP and GFP-ELP from that of the unfused GFP in transgenic soybean seeds based on the total soluble protein (TSP), despite the low-copy of T-DNA insertions and similar expression at the mRNA levels in selected transgenic lines. The average levels of zein-GFP and GFP-ELP accumulated in immature seeds of these transgenic lines were 0.99% and 0.29% TSP, respectively, compared with 0.07% TSP of the unfused GFP. In mature soybean seeds, the accumulation of zein-GFP and GFP-ELP proteins was 1.8% and 0.84% TSP, an increase of 3.91- and 1.82-fold, respectively, in comparison with that of the unfused GFP (0.46% TSP). Confocal laser scanning analysis showed that both zein-GFP and GFP-ELP were deposited in the protein storage vacuoles of transgenic seeds, while the unfused GFP was mainly distributed in the cytoplasm. Despite increased recombinant protein accumulation, there were no significant changes in the total protein and oil content in seeds between the transgenic and non-transformed plants, suggesting the possible presence of threshold limits of total protein accumulation in transgenic soybean seeds.
Conclusions
Overall, our results indicate that γ-zein and ELP fusions significantly increased the accumulation of the recombinant protein, but exhibited no significant influence on the total protein and oil content in soybean seeds.
Figure 1
Figure 2
Figure 3
Figure 4
Figure 5