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极化剂AsymPolPOK在完整的和溶解的哺乳动物细胞中的稳定性

Dominique Lagasca¹, Rupam Ghosh¹, Yiling Xiao¹

¹Department of Biophysics, UT Southwestern Medical Center, Dallas, TX 75390-8816.

bioRxiv : the preprint server for biology

|November 28, 2024

概括

一种新的极化剂,AsymPolPOK,显示出对细胞内动态核极化 (DNP) 的NMR有希望. 然而,其在细胞中的有效性受到快速减少的限制,因此需要进一步开发用于细胞结构生物学的生物耐药剂.

科学领域:

生物物理学的生物物理.
结构生物学结构生物学
生物化学生物化学

背景情况:

动态核极化 (DNP) 固态NMR增强了在内源水平检测蛋白质的灵敏度.
氧化物二基极化剂对于提高DNP灵敏度至关重要.
AsymPolPOK是一种新型,水溶性不对称的氧化物二基,具有提高DNP性能的潜力.

研究的目的:

为了描述哺乳动物HEK293细胞中的新型AsymPolPOK双基.
为了评估它的细胞分布,还原动力学和DNP性能.
评估其作为细胞内DNPNMR的极化剂的潜力.

主要方法:

电穿孔用于AsimPolPOK的细胞输送.
评估细胞分布和细胞毒性.
在细胞环境中减少AsymPolPOK的动力学研究.
在细胞溶解物和完整细胞中测量DNP增强和积累时间.

主要成果:

电穿孔使AsymPolPOK的细胞递送均,包括核透,没有观察到细胞毒性.
细胞环境迅速将AsymPolPOK降解为单基形式,其中氧化物中心之间的降解电阻差异.
AsymPolPOK在细胞溶解物中表现出高的DNP增强和快速积累时间.
在完整的细胞中,DNP的性能取决于输送方法,电穿孔的性能优于化,但随着长时间的细胞暴露,性能下降.

结论:

AsymPolPOK显示出作为细胞内DNPNMR的两极化剂的潜力.
细胞降低AsymPolPOK显著影响其在完整细胞中的DNP性能.
需要开发出更具生物耐药性的极化剂,以推进细胞内DNP NMR用于细胞结构生物学.

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