Max-Planck-Institut für Molekulare Pflanzenphysiologie, Am Mühlenberg 1, D-14476 Potsdam-Golm, Germany.
Plant Biotechnol J. 2011 Jun;9(5):599-608. doi: 10.1111/j.1467-7652.2011.00598.x. Epub 2011 Feb 11.
Plants with transgenic plastid (chloroplast) genomes represent a promising production platform in molecular farming, mainly because of the plastids' potential to accumulate foreign proteins to very high levels and the increased biosafety conferred by the maternal mode of plastid inheritance. Although some transgenes can be expressed to extraordinarily high levels, the expression of others has been unsuccessful. Lack of detectable transgene expression is usually attributable to either RNA instability or protein instability. Here, we have investigated the possibilities to improve the production of a pharmaceutical protein that is difficult to express in chloroplasts: the HIV-1 fusion inhibitor cyanovirin-N (CV-N). Testing various N-terminal and C-terminal fusions to peptide sequences from two proteins known to accumulate to high levels in transgenic plastids (GFP and the protein antibiotic PlyGBS), we show that both low mRNA stability and low protein stability contribute to the lack of detectable CV-N expression in chloroplasts. Both problems can be alleviated by N-terminal fusions to the CV-N coding region, thus highlighting a suitable strategy for optimization of plastid transgene expression.
具有转基因质体（叶绿体）基因组的植物在分子农业中代表了一个有前途的生产平台，主要是因为质体具有将外源蛋白积累到非常高水平的潜力，并且质体通过母系遗传方式赋予了增加的生物安全性。尽管一些转基因可以表达到极高的水平，但其他转基因的表达却不成功。缺乏可检测到的转基因表达通常归因于 RNA 不稳定性或蛋白质不稳定性。在这里，我们研究了提高一种在叶绿体中难以表达的药物蛋白产量的可能性：HIV-1 融合抑制剂氰基病毒-N（CV-N）。我们测试了各种与两种已知在转基因质体中积累到高水平的蛋白质（GFP 和蛋白抗生素 PlyGBS）的肽序列的 N 端和 C 端融合，结果表明低 mRNA 稳定性和低蛋白稳定性都导致叶绿体中无法检测到 CV-N 的表达。这两个问题都可以通过与 CV-N 编码区的 N 端融合来缓解，从而突出了优化质体转基因表达的合适策略。
