Open AccessInfluence of Elastin-Like Polypeptide and Hydrophobin on Recombinant Hemagglutinin Accumulations in Transgenic Tobacco Plants
Author(s) -
Hoang Trong Phan,
Bettina Hause,
Gerd Hause,
Elsa Arcalís,
Eva Stöger,
Daniel Maresch,
Friedrich Altmann,
Jussi Joensuu,
Udo Conrad
Publication year - 2014
Publication title -
plos one
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.99
H-Index - 332
ISSN - 1932-6203
DOI - 10.1371/journal.pone.0099347
Subject(s) - hydrophobin , kdel , hemagglutinin (influenza) , er retention , fusion protein , biology , endoplasmic reticulum , glycosylation , microbiology and biotechnology , recombinant dna , biochemistry , golgi apparatus , gene , mutant
Fusion protein strategies are useful tools to enhance expression and to support the development of purification technologies. The capacity of fusion protein strategies to enhance expression was explored in tobacco leaves and seeds. C-terminal fusion of elastin-like polypeptides (ELP) to influenza hemagglutinin under the control of either the constitutive CaMV 35S or the seed-specific USP promoter resulted in increased accumulation in both leaves and seeds compared to the unfused hemagglutinin. The addition of a hydrophobin to the C-terminal end of hemagglutinin did not significantly increase the expression level. We show here that, depending on the target protein, both hydrophobin fusion and ELPylation combined with endoplasmic reticulum (ER) targeting induced protein bodies in leaves as well as in seeds. The N -glycosylation pattern indicated that KDEL sequence-mediated retention of leaf-derived hemagglutinins and hemagglutinin-hydrophobin fusions were not completely retained in the ER. In contrast, hemagglutinin-ELP from leaves contained only the oligomannose form, suggesting complete ER retention. In seeds, ER retention seems to be nearly complete for all three constructs. An easy and scalable purification method for ELPylated proteins using membrane-based inverse transition cycling could be applied to both leaf- and seed-expressed hemagglutinins.