Department of Applied Genetics and Cell Biology, University of Natural Resources and Life Sciences , Vienna , Austria.
Institute of Molecular Biotechnology, RWTH Aachen University , Aachen , Germany.
Front Bioeng Biotechnol. 2014 Dec 11;2:67. doi: 10.3389/fbioe.2014.00067. eCollection 2014.
Naturally occurring storage proteins such as zeins are used as fusion partners for recombinant proteins because they induce the formation of ectopic storage organelles known as protein bodies (PBs) where the proteins are stabilized by intermolecular interactions and the formation of disulfide bonds. Endogenous PBs are derived from the endoplasmic reticulum (ER). Here, we have used different targeting sequences to determine whether ectopic PBs composed of the N-terminal portion of mature 27 kDa γ-zein added to a fluorescent protein could be induced to form elsewhere in the cell. The addition of a transit peptide for targeting to plastids causes PB formation in the stroma, whereas in the absence of any added targeting sequence PBs were typically associated with the plastid envelope, revealing the presence of a cryptic plastid-targeting signal within the γ-zein cysteine-rich domain. The subcellular localization of the PBs influences their morphology and the solubility of the stored recombinant fusion protein. Our results indicate that the biogenesis and budding of PBs does not require ER-specific factors and therefore, confirm that γ-zein is a versatile fusion partner for recombinant proteins offering unique opportunities for the accumulation and bioencapsulation of recombinant proteins in different subcellular compartments.
天然存在的贮藏蛋白，如 zein，被用作重组蛋白的融合伴侣，因为它们诱导形成异位贮藏细胞器，称为蛋白体（PBs），在这些细胞器中，蛋白质通过分子间相互作用和形成二硫键而稳定。内源性 PBs 来源于内质网（ER）。在这里，我们使用了不同的靶向序列来确定添加到荧光蛋白的成熟 27 kDa γ-zein 的 N 端部分组成的异位 PBs 是否可以在细胞的其他部位诱导形成。添加靶向质体的转运肽会导致 PB 在基质中形成，而在没有添加任何靶向序列的情况下，PB 通常与质体包膜相关，这表明 γ-zein 半胱氨酸丰富结构域内存在隐蔽的质体靶向信号。PB 的亚细胞定位影响其形态和储存重组融合蛋白的溶解度。我们的结果表明，PB 的生物发生和出芽不需要 ER 特异性因子，因此证实 γ-zein 是重组蛋白的多功能融合伴侣，为重组蛋白在不同亚细胞区室中的积累和生物封装提供了独特的机会。
