Bellucci Michele, De Marchis Francesca, Nicoletti Ildo, Arcioni Sergio
Institute of Plant Genetics - Research Division of Perugia, Italian National Research Council (CNR), via della Madonna Alta 130, 06128 Perugia, Italy.
J Biotechnol. 2007 Aug 31;131(2):97-105. doi: 10.1016/j.jbiotec.2007.06.004. Epub 2007 Jun 19.
Several strategies have been exploited to maximize heterologous protein accumulation in the plant cell. Recently, it has been shown that a portion of a maize prolamin storage protein, gamma-zein, can be used for the high accumulation of a recombinant protein in novel endoplasmic reticulum (ER)-derived protein bodies of vegetative tissues. In this study, we investigate whether this protein can be expressed in the chloroplast. Our long-term purpose is to use zeolin to produce value-added proteins by fusing these polypeptides with its gamma-zein portion and targeting the recombinant proteins to the ER or to the chloroplast. We show here that zeolin accumulates in the chloroplast to lower levels than in the ER and its stability is compromised by chloroplast proteolytic activity. Co-localization of zeolin and the ER chaperone BiP in the chloroplast does not have a beneficial effect on zeolin accumulation. In this organelle, zeolin is not stored in protein bodies, nor do zeolin polypeptides seem to be linked by inter-chain disulfide bonds, which are usually formed by the six cysteine of the gamma-zein portion, indicating abnormal folding of the recombinant protein. Therefore, it is concluded that to accumulate zeolin in the chloroplast it is necessary to facilitate inter-chain disulfide bond formation.
人们已经采用了多种策略来使植物细胞中的异源蛋白积累最大化。最近有研究表明，玉米醇溶蛋白储存蛋白γ-玉米醇溶蛋白的一部分可用于在营养组织中新形成的内质网（ER）衍生蛋白体中高效积累重组蛋白。在本研究中，我们探究了这种蛋白是否能在叶绿体中表达。我们的长期目标是通过将这些多肽与γ-玉米醇溶蛋白部分融合，并将重组蛋白靶向到内质网或叶绿体，利用玉米醇溶蛋白来生产增值蛋白。我们在此表明，玉米醇溶蛋白在叶绿体中的积累水平低于在内质网中的积累水平，并且其稳定性受到叶绿体蛋白水解活性的影响。玉米醇溶蛋白与内质网伴侣BiP在叶绿体中的共定位对玉米醇溶蛋白的积累没有有益影响。在这个细胞器中，玉米醇溶蛋白不储存在蛋白体中，玉米醇溶蛋白多肽似乎也没有通过链间二硫键连接，而链间二硫键通常由γ-玉米醇溶蛋白部分的六个半胱氨酸形成，这表明重组蛋白折叠异常。因此，得出的结论是，要在叶绿体中积累玉米醇溶蛋白，有必要促进链间二硫键的形成。
