School of Veterinary Medicine, University of Pennsylvania, New Bolton Center Campus, 382 West Street Road, Kennett Square, PA 19348, USA.
J Am Soc Mass Spectrom. 2011 Apr;22(4):718-30. doi: 10.1007/s13361-011-0080-5. Epub 2011 Feb 18.
Identification of an unknown substance without any information remains a daunting challenge despite advances in chemistry and mass spectrometry. However, an unknown cyclic peptide in a sample with very limited volume seized at a Pennsylvania racetrack has been successfully identified. The unknown sample was determined by accurate mass measurements to contain a small unknown peptide as the major component. Collision-induced dissociation (CID) of the unknown peptide revealed the presence of Lys (not Gln, by accurate mass), Phe, and Arg residues, and absence of any y-type product ion. The latter, together with the tryptic digestion results of the unusual deamidation and absence of any tryptic cleavage, suggests a cyclic structure for the peptide. Electron-transfer dissociation (ETD) of the unknown peptide indicated the presence of Gln (not Lys, by the unusual deamidation), Phe, and Arg residues and their connectivity. After all the results were pieced together, a cyclic tetrapeptide, cyclo[Arg-Lys-N(C(6)H(9))Gln-Phe], is proposed for the unknown peptide. Observations of different amino acid residues from CID and ETD experiments for the peptide were interpreted by a fragmentation pathway proposed, as was preferential CID loss of a Lys residue from the peptide. ETD was used for the first time in sequencing of a cyclic peptide; product ions resulting from ETD of the peptide identified were categorized into two types and named pseudo-b and pseudo-z ions that are important for sequencing of cyclic peptides. The ETD product ions were interpreted by fragmentation pathways proposed. Additionally, multi-stage CID mass spectrometry cannot provide complete sequence information for cyclic peptides containing adjacent Arg and Lys residues. The identified cyclic peptide has not been documented in the literature, its pharmacological effects are unknown, but it might be a "designer" drug with athletic performance-enhancing effects.
尽管化学和质谱技术取得了进步，但对于没有任何信息的未知物质的鉴定仍然是一个艰巨的挑战。然而，在宾夕法尼亚州赛马场缴获的一个体积非常有限的样本中，一种未知的环状肽已经被成功鉴定。通过精确质量测量，确定未知样品的主要成分是一个小的未知肽。对未知肽的碰撞诱导解离（CID）揭示了存在赖氨酸（不是精氨酸，根据精确质量）、苯丙氨酸和精氨酸残基，并且没有任何 y 型产物离子。后一种情况，加上胰蛋白酶消化的不寻常脱酰胺和没有任何胰蛋白酶切割的结果，表明该肽具有环状结构。对未知肽的电子转移解离（ETD）表明存在谷氨酰胺（不是赖氨酸，通过不寻常的脱酰胺）、苯丙氨酸和精氨酸残基及其连接性。将所有结果拼凑在一起后，提出了一种未知肽的环状四肽，cyclo[Arg-Lys-N(C(6)H(9))Gln-Phe]。通过提出的碎裂途径解释了 CID 和 ETD 实验中观察到的不同氨基酸残基，以及肽中优先从赖氨酸残基发生 CID 丢失。ETD 首次用于环状肽的测序；肽的 ETD 产生的产物离子被分为两类，并分别命名为 pseudo-b 和 pseudo-z 离子，它们对环状肽的测序很重要。通过提出的碎裂途径解释了 ETD 产物离子。此外，多阶段 CID 质谱不能为含有相邻精氨酸和赖氨酸残基的环状肽提供完整的序列信息。所鉴定的环状肽在文献中没有记载，其药理作用未知，但它可能是一种具有增强运动表现效果的“设计药物”。
