反相高效液相色谱-质谱联用法测定抑菌肽功能单元的氨基酸序列

Determination of Amino Acid Sequence of Antibacterial Peptide Function Units by Using RPHPLC-MS Method

DOI：10.3969/j.issn.1673-1689.2014.05.012

中文关键词: 小分子抑菌肽 凝胶过滤色谱 反相高效液相色谱 二维质谱 氨基酸序列

英文关键词: small antibacterial peptide, gel-filtration chromatography, reversed-phase high performance liquid chromatography, two-dimensional mass spectrometry, amino acid sequence

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中文摘要:

测定乳酸菌发酵液中小分子抑菌肽抑菌功能单元的氨基酸组成序列对于深入理解其抑菌原理并实际应用有重要意义。作者利用AssayMAP Bravo蛋白纯化系统平台，采用色谱-质谱联用法将相对分子质量为3 500的抑菌肽样品经过凝胶过滤色谱分离纯化，两步反相高效液相色谱分离纯化和二维质谱鉴定，得到高纯度、高抑菌活性的三个抑菌功能单元，氨基酸组成序列分别为Phe-Tyr-Pro-Ser-Tyr-Ala，Asn-Glu-Arg-His和Lys-Glu-Ile-Thr-Pro-Ser-Glu-Arg，即该小肽的功能单元由三个短肽组成。结果表明，利用该方法，能够有效地提取并检测该小分子抑菌肽，其在氨基酸组成上与Nisin不同。

英文摘要:

The objective of this study is to determine the amino acid sequence of antibacterial peptide function units from Lacticacid bacteria fermented liquid, it has important significance for deep understanding of the antibacterial principle and practical application.According to the AssayMAP Bravo protein purification system, the experiment used the RPHPLC-MS method, got three components of high purity and high antibacterial activity from the 3.5 KDa small antibacterial peptide samples by using Gel-Filtration Chromatography, two-steps Reversed-Phase High Performance Liquid Chromatography and two-dimensional Mass Spectrometry. The amino acid sequence of three antibacterial peptide function units had been measured, respectively were Phe-Tyr-Pro-Ser-Tyr-Ala, Asn-Glu-Arg-His and Lys-Glu-Ile-Thr-Pro-Ser-Glu-Arg. The experimental conclusion is that the antibacterial function units of small peptide consists of three short peptides, using this method can effectively extract and detection of the small antibacterial peptide, and the amino acid composition of 3.5 KDa small antibacterial peptide is different from Nisin.

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