Smart Protein Refolding System Based on UCST-Type Ureido Polymers
Smart Protein Refolding System Based on UCST-Type Ureido Polymers
复制标题
基于UCST型Ureido聚合物的智能蛋白质重折叠系统
DOI：
10.1021/acs.biomac.2c00694
复制
发表时间：
2022
影响因子：
6.2
通讯作者：
Shimada Naohiko
中科院分区：
文献类型：
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作者：
Tanaka Yamato;Niu Chun Hao;Sasaki Taira;Nomura Shouhei;Maruyama Atsushi;Shimada Naohiko
我们已经报道了脲基聚合物在溶液中表现出上临界溶解温度（UCST）型相行为，这与下临界溶解温度（LCST）型行为相反。此外，UCST型脲基聚合物在冷却时经历液-液相分离（LLPS），而不是典型LCST型聚合物的液-固相转变。在这项研究中，制备了具有疏水基团的脲基聚合物，以评估冷却诱导的UCST型聚合物的LLPS对蛋白质复性的影响。当蛋白质与用十一烷基官能化的脲基聚合物一起加热时，蛋白质的聚集被阻止。随后的冷却孵育导致蛋白质从聚合物中自发释放。释放的蛋白质具有酶活性，表明蛋白质正确地重折叠。有趣的是，当UCST型脲基聚合物和蛋白质的溶液在聚合物的相分离温度附近孵育时，观察到有效的重折叠，这意味着聚合物的冷却诱导的LLPS增强了蛋白质的释放。另外，通过在4 °C下离心，重折叠的蛋白质容易与脲基聚合物分离，脲基聚合物在冷却时沉淀。
We have reported that ureido polymers exhibit upper critical solution temperature (UCST)-type phase behavior in solution, which is the opposite of lower critical solution temperature (LCST)-type behavior. Furthermore, UCST-type ureido polymers undergo liquid–liquid phase separation (LLPS) upon cooling rather than the liquid–solid phase transition of the typical LCST-type polymers. In this study, ureido polymers with hydrophobic groups were prepared to evaluate the effects of cooling-induced LLPS of UCST-type polymers on refolding of proteins. When protein was heated with a ureido polymer functionalized with undecyl groups, aggregation of the protein was prevented. Subsequent cooling incubation resulted in the spontaneous release of the protein from the polymer. The released protein had enzymatic activity, suggesting that the protein refolded properly. Interestingly, efficient refolding was observed when the solution of the UCST-type ureido polymer and protein was incubated at around the phase separation temperature of the polymer, implying that cooling-induced LLPS of the polymer enhanced the release of the protein. Additionally, by centrifugation at 4 °C, the refolded protein was readily separated from the ureido polymers, which precipitated upon cooling.