剪切诱导的结构变化驱动人类胰岛素的无形聚合物形成
Chinmaya Panda1, Sachin Kumar2, Sharad Gupta3
1Bio-Interface & Environmental Engineering Lab, Department of Biosciences and Bioengineering, Indian Institute of Technology Guwahati, Assam, India.
Proteins
|June 30, 2025
概括
剪切应力,加热和低pH值,通过促进展开和纤维细胞形成,显著加快人体胰岛素聚合. 控制这些因素对于生物制药的稳定性至关重要.
科学领域:
- 生物制药科学 生物制药科学
- 蛋白质化学 蛋白质化学
- 类风病学 类风病学 类风病学
背景情况:
- 蛋白质聚合是生物制药配方的主要挑战,影响稳定性和疗效.
- 人类胰岛素 (HI) 聚合在pH和温度变化下得到了很好的研究,但联合压力因素的理解较少.
- 温度,pH和剪切等物理化学压力因素可以协同破坏蛋白质的完整性.
研究的目的:
- 研究人类胰岛素 (HI) 在pH,剪切和热应力组合下的聚合动力学.
- 阐明由这些压力因素引起的形状变化和聚合机制.
- 评估剪切诱导聚合对HI的结构完整性和细胞毒性的影响.
主要方法:
- 在不同的剪速 (1-1000 s-1) 和温度 (25-60°C) 中评估HI聚合动力学.
- 使用了度法 (Thioflavin-T,内在的氨酸光),循环二极化和传输电子显微镜 (TEM).
- 采用原生PAGE,BCA测定单体耗尽,和细胞毒性研究.
主要成果:
- 在60°C和低pH下,HI表现出非牛顿式的行为,剪切变厚,随后由于聚合物碎片化而变薄.
- 剪切诱导的能量超过了展开的自由能量,催化了展开,β片形成和聚合.
- 剪切减少了80倍的纤维化滞后时间;在高剪切和高温下,α-螺旋体含量显著下降;TEM显示了纤维化到无形的过渡.
结论:
- 剪切应力作为人类胰岛素聚合的强有力的催化剂,加剧了展开和纤维的形成.
- 结合的热力学压力因素显著影响HI的结构完整性,并可能导致细胞活力降低.
- 在胰岛素生物处理,运输和储存过程中,严格控制温度和切割是必不可少的,以确保生物制药的稳定性.
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