混合/-核酸结构的完整测序使用MALDI-TOF/TOF质谱仪
Daniil G Ivanov1, John J Thomas2, Igor A Kaltashov1
1Department of Chemistry, University of Massachusetts-Amherst, Amherst, Massachusetts 01003, United States.
Analytical chemistry
|November 5, 2025
概括
本研究引入了一种质谱法,用于测序复杂的核酸 (PNA). 这种技术可以进行详细的结构特征,这对于开发先进的PNA疗法至关重要.
科学领域:
- 生物化学 生物化学
- 分析化学 分析化学
- 药用化学 医学化学
背景情况:
- 核酸 (PNA) 显示出治疗前景,但与其他核酸类似物相比,在临床应用中面临挑战.
- 最近PNA设计的进展涉及复杂的结构,其中包括脊柱修改和片段,以提高稳定性,特异性和传递性.
- 鉴定这些大而复杂的PNA分子是很困难的,因为它们独特的结构特征和非自然的脊柱.
研究的目的:
- 开发和验证一个自上而下的MS/MS方法,用于新的复杂的新一代PNA分子的新序列和结构特征.
- 克服传统碎片化方法和酶分析在表征具有非自然脊椎和改性结构的PNAs方面的局限性.
- 为了能够精确地定位骨干修改,如 γ-基甲基,并区分异构PNA序列.
主要方法:
- 采用了自上而下的质谱/质谱 (MS/MS) 方法,特别是MALDI TOF/TOF,用于分析6kDa混合PNA分子.
- 采用高能碰撞激活来诱导整个PNA分子的碎片化,包括非天然的骨干.
- 开发了一种使用碎片离子光谱来区分异构PNA序列的"质量指纹"策略.
主要成果:
- 实现了混合PNA分子的完整序列覆盖,包括所有γ-基甲基修饰的局部化.
- 证明了 de novo 测序复杂的 PNA 结构的能力,克服了传统方法中电荷定位的局限性.
- 通过使用质量指纹,成功地区分了参考PNA和具有编码序列的异构分子.
结论:
- 在MALDI TOF/TOF MS/MS中高能碰撞激活对于复杂的PNA分子的全面结构特征是有效的.
- 这种基于MS的方法促进了先进PNA疗法的de novo测序和精确的结构分析.
- 开发的大规模指纹技术为基于PNA的产品的质量控制和序列验证提供了强大的方法.
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