重新审视三素消化缓冲剂对人工除菌的作用
Emmajay Sutherland1, Tim S Veth1, Nicholas M Riley1
1Department of Chemistry, University of Washington, Seattle, Washington 98195, United States.
Journal of the American Society for Mass Spectrometry
|January 31, 2025
概括
通过选择适当的缓冲区,可以将蛋白质组学中的化学除化最小化. HEPES缓冲体显示出最少的人工除化,使其适合蛋白质组学研究,特别是那些涉及N-糖化化的研究.
科学领域:
- 蛋白质组学是指蛋白质组学.
- 生物化学 生物化学
- 分析化学 分析化学
背景情况:
- 阿斯巴拉金和谷氨胺残留物的脱化是一种常见的翻译后修饰.
- 在自下而上的蛋白质组学中,蛋白质溶解消化可以诱导人造的脱氧化,从而影响实验结果.
- 尽量减少化学除化对于准确的蛋白质组学分析至关重要.
研究的目的:
- 重新评估和比较缓冲系统,以最大限度地减少化学诱导的蛋白质组学中的脱化.
- 评估生物相容的缓冲剂,包括HEPES,EPPS和PBS,是否适用于减少除化.
主要方法:
- 标准的自下而上的蛋白质组学工作流程.
- 对Tris-HCl,二碳酸,三乙二碳酸,HEPES,EPPS和PBS缓冲剂的评估.
- 在特定的化条件下,人工除化量的量化.
主要成果:
- 与其他测试的缓冲器相比,HEPES缓冲器显示了最少的人工除化.
- 酸,虽然以前建议,提出了与素消化和测试的兼容性问题.
- 像HEPES这样的生物相容缓冲剂是最小化除化的一种可行的选择.
结论:
- 对于一般的蛋白质组实验来说,HEPES缓冲器是推的选择,以最大限度地减少人工除化.
- 缓冲系统的选择显著影响了蛋白质学中的化学除化程度.
- 这一发现对于涉及N-糖化酶的研究尤为重要,在这些研究中,脱胺化可以干扰分析.
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