Nausch Henrik, Dorn Mandy, Frolov Andrej, Hoedtke Sandra, Wolf Petra, Broer Inge
Department of Agrobiotechnology and Risk Assessment for Bio- und Gene Technology, Faculty of Agricultural and Environmental Sciences, University of Rostock, Rostock, Germany.
Department of Bioorganic Chemistry, Leibniz Institute of Plant Biochemistry, Halle (Saale), Germany.
Front Plant Sci. 2020 Jun 19;11:842. doi: 10.3389/fpls.2020.00842. eCollection 2020.
Feed supplementation with β-arginine-aspartate dipeptides (β-Asp-Arg DP) shows growth promoting effects in feeding trials with fish and might also be beneficial for pig and poultry farming. Currently, these DPs are generated from purified cyanophycin (CGP), with the help of the CGP-degrading enzyme cyanophycinase (CGPase). As alternative to an production, the DPs might be directly produced in feed crops. We already demonstrated that CGP can be produced in plastids of tobacco and potato, yielding up to 9.4% of the dry weight (DW). We also transiently co-expressed CGPases in the cytosol without degrading CGP in intact cells, while degradation occurs in the homogenized plant tissue. However, transient co-expression is not feasible for field-grown CGP plants, which is necessary for bulk production. In the present study, we proved that stable co-expression of the CGPase CphE241 in CGP-producing tobacco is sufficient to degrade 2.0% CGP/DW nearly completely within 3 h after homogenization of the leaves. In intact senescing leaves, CGP is partially released to the cytosol and degraded into DPs which limits the overall accumulation of CGP but not the level of the stable DPs. Even after 48 h, 54 μmol β-Asp-Arg DP/g DW could be detected in the extract, which correspond to 1.5% DP/DW and represents 84% of the expected amount. Thus, we developed a system for the production of β-Asp-Arg DP in field-grown plants.
在鱼类饲养试验中，添加β-精氨酸-天冬氨酸二肽（β-Asp-Arg DP）的饲料显示出促进生长的作用，对养猪和家禽养殖可能也有益处。目前，这些二肽是借助于降解藻青素（CGP）的酶——藻青素酶（CGPase）从纯化的藻青素中生成的。作为一种替代生产方式，二肽或许可以在饲料作物中直接产生。我们已经证明，CGP能够在烟草和马铃薯的质体中产生，产量高达干重的9.4%。我们还在细胞质中瞬时共表达了CGPase，但在完整细胞中并未降解CGP，而在匀浆的植物组织中则会发生降解。然而，对于田间种植的用于大量生产CGP的植物而言，瞬时共表达是不可行的。在本研究中，我们证明在生产CGP的烟草中稳定共表达CGPase CphE241足以在叶片匀浆后3小时内将2.0%的CGP/DW几乎完全降解。在完整的衰老叶片中，CGP会部分释放到细胞质中并降解为二肽，这限制了CGP的总体积累，但不会影响稳定二肽的水平。即便在48小时后，提取物中仍可检测到54 μmol β-Asp-Arg DP/g DW，这相当于1.5%的DP/DW，占预期量的84%。因此，我们开发了一种在田间种植的植物中生产β-Asp-Arg DP的系统。
