Protein body formation in stable transgenic tobacco expressing elastin-like polypeptide and hydrophobin fusion proteinsReport as inadecuate




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BMC Biotechnology

, 13:40

Plant biotechnology

Abstract

BackgroundPlants are recognized as an efficient and inexpensive system to produce valuable recombinant proteins. Two different strategies have been commonly used for the expression of recombinant proteins in plants: transient expression mediated by Agrobacterium; or stable transformation of the plant genome. However, the use of plants as bioreactors still faces two main limitations: low accumulation levels of some recombinant proteins and lack of efficient purification methods. Elastin-like polypeptide ELP, hydrophobin I HFBI and Zera® are three fusion partners found to increase the accumulation levels of recombinant proteins and induce the formation of protein bodies PBs in leaves when targeted to the endoplasmic reticulum ER in transient expression assays. In this study the effects of ELP and HFBI fusion tags on recombinant protein accumulation levels and PB formation was examined in stable transgenic Nicotiana tabacum.

ResultsThe accumulation of recombinant protein and PB formation was evaluated in two cultivars of Nicotiana tabacum transformed with green fluorescent protein GFP fused to ELP or HFBI, both targeted and retrieved to the ER. The ELP and HFBI tags increased the accumulation of the recombinant protein and induced the formation of PBs in leaves of stable transgenic plants from both cultivars. Furthermore, these tags induced the formation of PBs in a concentration-dependent manner, where a specific level of recombinant protein accumulation was required for PBs to appear. Moreover, agro-infiltration of plants accumulating low levels of recombinant protein with p19, a suppressor of post-transcriptional gene silencing PTGS, increased accumulation levels in four independent transgenic lines, suggesting that PTGS might have caused the low accumulation levels in these plants.

ConclusionThe use of ELP and HFBI tags as fusion partners in stable transgenic plants of tobacco is feasible and promising. In a constitutive environment, these tags increase the accumulation levels of the recombinant protein and induce the formation of PBs regardless of the cultivar used. However, a specific level of recombinant protein accumulation needs to be reached for PBs to form.

KeywordsProtein body Protein body formation Elastin-like polypeptide ELP Hydrophobin I HFBI Tobacco Transgenic expression Molecular farming Green fluorescent protein AbbreviationsATPSAqueous two-phase system

CymRSVCymbidium ringspot tombusvirus

ELPElastin-like polypeptide

GFPGreen fluorescent protein

HFBIHydrophobin I

ITCInverse transition cycling

PBProtein body

PTGSPost-transcriptional gene silencing

TSPTotal soluble protein.

Electronic supplementary materialThe online version of this article doi:10.1186-1472-6750-13-40 contains supplementary material, which is available to authorized users.

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Author: Sonia P Gutiérrez - Reza Saberianfar - Susanne E Kohalmi - Rima Menassa

Source: https://link.springer.com/







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