Efficient synthesis of amino acid polymers for protein stabilization
Efficient synthesis of amino acid polymers for protein stabilization
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高效合成用于稳定蛋白质的氨基酸聚合物
DOI：
10.1039/c9bm00484j
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发表时间：
2019-09-01
影响因子：
6.6
通讯作者：
Liu, Runhui
中科院分区：
文献类型：
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作者：
Li, Bing;Wu, Yueming;Liu, Runhui
蛋白质是易碎的，即使冷冻、干燥和脱水也可能导致其变性、聚集和活性丧失。为了保护蛋白质免受这些类型的损伤，我们制备了两种类型的氨基酸聚合物，聚-（L-谷氨酸）-r-聚-（L-赖氨酸）（PLG-r-PLL）和聚-L-谷氨酸（PLG），从高效开环聚合的α-氨基酸N-羧酸酐（NCA）使用六甲基二硅氮基锂（LiHMDS）作为引发剂。β-半乳糖苷酶（β-Gal）用于本研究以检查在冻干过程中合成的氨基酸聚合物的蛋白质保护效果。结果表明，PLG-r-PLL和PLG在冻干过程中对β-Gal都发挥了显著的保护作用，并将所得蛋白质的活性从冻干过程中不使用保护剂的40%提高到原始蛋白质活性的80%。然而，PLG通常比PLG-r-PLL表现得更好，与链长度无关。我们的研究还表明，PLG和PLG-r-PLL具有高含量的PLG亚基显示没有观察到的细胞毒性和溶血作用。此外，动态光散射（DLS）和透射电子显微镜（TEM）表征表明，PLG通过防止β-Gal的聚集在冻干时保护β-Gal。我们的研究表明，氨基酸聚合物，如PLG，可以发挥有效的活性，蛋白质稳定。LiHMDS引发的NCA聚合反应操作简单，反应效率高，这表明该方法在快速制备氨基酸聚合物方面具有很大的潜力，可用于蛋白质稳定性筛选和机理研究。
Proteins are fragile such that even freezing, drying and dehydration may induce their denaturation, aggregation, and activity loss. To protect proteins from these kinds of damage, we prepared two types of amino acid polymers, poly-(l-glutamate)-r-poly-(l-lysine) (PLG-r-PLL) and poly-l-glutamate (PLG), from the efficient ring-opening polymerization of alpha-amino acid N-carboxyanhydride (NCA) using lithium hexamethyldisilazide (LiHMDS) as the initiator. beta-galactosidase (beta-Gal) was used in this study to examine the protein protecting effect of the synthesized amino acid polymers during lyophilization. The results indicate that both PLG-r-PLL and PLG exert significant protection on beta-Gal during lyophilization and improve the activity of the resulting protein from 40%, without using a protecting agent during lyophilization, to 80% of the original protein activity. Nevertheless, PLG generally performs better than PLG-r-PLL independent of the chain length. Our studies also show that PLG and PLG-r-PLL with a high content of PLG subunits display no observable cytotoxicity and hemolytic effect. Furthermore, dynamic light scattering (DLS) and transmission electron microscopy (TEM) characterization indicate that PLG protects beta-Gal upon lyophilization by preventing the aggregation of beta-Gal. Our studies demonstrate that amino acid polymers, such as PLG, can exert potent activity for protein stabilization. The easy operation of LiHMDS-initiated and efficient NCA polymerization implies the great potential of this strategy to prepare amino acid polymers quickly for the screening of protein stabilization and mechanism study.