Molecular Farming in Barley: Development of a Novel Production Platform to Produce Human Antimicrobial Peptide LL‐37,Biotechnology Journal

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Molecular Farming in Barley: Development of a Novel Production Platform to Produce Human Antimicrobial Peptide LL‐37
Biotechnology Journal ( IF 3.2 ) Pub Date : 2018-02-08 , DOI: 10.1002/biot.201700628
Edita Holásková ₁ , Petr Galuszka ₁ , Alžbeta Mičúchová ₁ , Marek Šebela ₁ , Mehmet Tufan Öz ₁ , Ivo Frébort ₁

Affiliation

The peptide LL‐37, a component of the human innate immune system, represents a promising drug candidate. In particular, the development of low‐cost production platform technology is a critical bottleneck in its use in medicine. In the present study, a viable approach for the LL‐37 production in transgenic barley is developed. First, comparative analyses of the effects of different fused peptide epitope tags applicable for accumulation and purification on LL‐37 production yield are performed using transient expression in tobacco leaves. Following the selection of the most yielding fusion peptide strategies, eight different constructs for the expression of codon optimized chimeric LL‐37 genes in transgenic barley plants are created. The expression of individual constructs is driven either by an endosperm‐specific promoter of the barley B1 hordein gene or by the maize ubiquitin promoter. The transgenes are stably integrated into the barley genome and inherited in the subsequent generation. All transgenic lines show normal phenotypes and are fertile. LL‐37 accumulated in the barley seeds up to 0.55 mg per 1 kg of grain. The fused epitope tags are cleaved off by the use of enterokinase. Furthermore, in planta produced LL‐37 including the fused versions is biologically active.

中文翻译：

大麦的分子农业：开发新型生产平台来生产人类抗菌肽LL-37

LL-37肽是人类先天免疫系统的组成部分，代表着一种有前途的候选药物。特别是，低成本生产平台技术的发展是其在医学中使用的关键瓶颈。在本研究中，开发了一种在转基因大麦中生产LL-37的可行方法。首先，使用烟草叶片中的瞬时表达，对适用于积累和纯化的不同融合肽表位标签对LL-37产量的影响进行了比较分析。在选择了产量最高的融合肽策略后，用于密码子优化的嵌合LL-37表达的八种不同构建体在转基因大麦植物中创建基因。大麦B1大麦醇溶蛋白基因的胚乳特异性启动子或玉米泛素启动子驱动单个构建体的表达。转基因被稳定地整合到大麦基因组中并在后代中遗传。所有转基因品系均显示正常表型且可育。大麦种子中积累的LL-37达每1千克谷物0.55毫克。通过使用肠激酶将融合的表位标签切割掉。此外，在植物中生产的LL‐37（包括融合版本）具有生物活性。

更新日期：2018-06-03

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