A transgenic protein is frequently expressed as different homologous variants in genetically modified crops due to differential processing of targeting peptides or optimization of activity and specificity. The aim of this study was to develop a science-based approach for risk assessment of homologous protein variants using dicamba mono-oxygenase (DMO) as a case study. In this study, DMO expressed in the next-generation dicamba-tolerant maize, sugar beet and soybean crops exhibited up to 27 amino acid sequence differences in the N-terminus. Structure modeling using AlphaFold, ESMFold and OpenFold demonstrates that these small N-terminal extensions lack an ordered secondary structure and do not disrupt the DMO functional structure. Three DMO variants were demonstrated to have equivalent immunoreactivity and functional activity ranging from 214 to 331 nmol/min/mg. Repeated toxicity studies using each DMO variant found no test substance-related adverse effects. These results support that homologous protein variants, which have demonstrated physicochemical and functional equivalence, can leverage existing safety data from one variant without requiring additional de novo safety assessments.

中文翻译：

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使用桥接方法对生物技术性状蛋白的同源变体进行风险评估。


由于靶向肽的差异加工或活性和特异性的优化，转基因蛋白在转基因作物中经常表达为不同的同源变体。本研究的目的是开发一种基于科学的方法，以麦草畏单加氧酶 （DMO） 为案例研究，对同源蛋白质变体进行风险评估。在本研究中，在下一代耐麦草畏玉米、甜菜和大豆作物中表达的 DMO 在 N 端表现出多达 27 个氨基酸序列差异。使用 AlphaFold、ESMFold 和 OpenFold 的结构建模表明，这些小的 N 端延伸缺乏有序的二级结构，并且不会破坏 DMO 功能结构。三种 DMO 变体被证明具有等效的免疫反应性和功能活性，范围为 214 至 331 nmol/min/mg。使用每种 DMO 变体的重复毒性研究未发现与受试物质相关的不良反应。这些结果支持，已证明物理化学和功能等效性的同源蛋白质变体可以利用来自一个变体的现有安全性数据，而无需额外的从头安全性评估。