n端二甲基标记法测定蛋白质n端氨基酸序列:与Edman降解序列分析相比的缺陷与优势

Elizabeth Chang, S. Pourmal, Chun Zhou, Rupesh Kumar, M. Teplova, N. Pavletich, K. Marians, H. Erdjument-Bromage
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引用次数: 10

摘要

在最近的历史中,已经研究了Edman测序的替代方法,为此,生物分子资源设施协会(ABRF)蛋白质测序研究小组(PSRG)于2014年和2015年启动了研究,研究标准数量完整蛋白质的自下而上和自上而下n端(Nt)二甲基衍生化,目的是确定Nt序列信息。我们已经扩大了这一举措,并使用低皮摩尔量的肌红蛋白来确定nt -二甲基化的效率。这种方法在蛋白质结构域上的应用,由过表达蛋白的有限蛋白水解产生,证实了它是一种通用的标记技术,与Edman测序相比非常敏感。最后,我们比较了相同多肽片段的Edman测序和nt -二甲基化;结果证实两种方法鉴定的Nt氨基酸序列一致。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
N-Terminal Amino Acid Sequence Determination of Proteins by N-Terminal Dimethyl Labeling: Pitfalls and Advantages When Compared with Edman Degradation Sequence Analysis.
In recent history, alternative approaches to Edman sequencing have been investigated, and to this end, the Association of Biomolecular Resource Facilities (ABRF) Protein Sequencing Research Group (PSRG) initiated studies in 2014 and 2015, looking into bottom-up and top-down N-terminal (Nt) dimethyl derivatization of standard quantities of intact proteins with the aim to determine Nt sequence information. We have expanded this initiative and used low picomole amounts of myoglobin to determine the efficiency of Nt-dimethylation. Application of this approach on protein domains, generated by limited proteolysis of overexpressed proteins, confirms that it is a universal labeling technique and is very sensitive when compared with Edman sequencing. Finally, we compared Edman sequencing and Nt-dimethylation of the same polypeptide fragments; results confirm that there is agreement in the identity of the Nt amino acid sequence between these 2 methods.
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