西湖大学朱听团队报道了镜像胰蛋白酶消化和D蛋白测序。相关研究成果发表在2024年1月18日出版的国际知名学术期刊《自然—化学》。

镜像生物学系统及其相关应用的发展，由于缺乏对镜像（D-）蛋白进行测序的有效方法而受到阻碍。尽管天然手性（L-）蛋白可以通过自下而上的液相色谱-串联质谱法（LC–MS/MS）进行测序，但使用相同策略对长D肽和D蛋白进行测序需要在质量分析前，通过位点特异性D蛋白酶进行消化。

该文中，研究人员应用固相肽合成和天然化学连接来化学合成镜像版本的胰蛋白酶，这是一种广泛用于位点特异性蛋白质消化的蛋白酶。使用镜像胰蛋白酶消化和LC–MS/MS，研究人员对镜像大亚基核糖体蛋白（L25）和镜像溶脂硫酸藻P2 DNA聚合酶IV（Dpo4）进行测序，并区分D-Dpo4的不同突变体。研究人员还对50个氨基酸的长D肽中的数字信息进行写入和读取。因此，镜像胰蛋白酶消化结合LC–MS/MS可能有助于D肽和D蛋白，作为潜在的治疗和信息工具的实际应用。

附：英文原文

Title: Mirror-image trypsin digestion and sequencing of D-proteins

Author: Zhang, Guanwei, Zhu, Ting F.

Issue&Volume: 2024-01-18

Abstract: The development of mirror-image biology systems and related applications is hindered by the lack of effective methods to sequence mirror-image (D-) proteins. Although natural-chirality (L-) proteins can be sequenced by bottom–up liquid chromatography–tandem mass spectrometry (LC–MS/MS), the sequencing of long D-peptides and D-proteins with the same strategy requires digestion by a site-specific D-protease before mass analysis. Here we apply solid-phase peptide synthesis and native chemical ligation to chemically synthesize a mirror-image version of trypsin, a widely used protease for site-specific protein digestion. Using mirror-image trypsin digestion and LC–MS/MS, we sequence a mirror-image large subunit ribosomal protein (L25) and a mirror-image Sulfolobus solfataricus P2 DNA polymerase IV (Dpo4), and distinguish between different mutants of D-Dpo4. We also perform writing and reading of digital information in a long D-peptide of 50 amino acids. Thus, mirror-image trypsin digestion in conjunction with LC–MS/MS may facilitate practical applications of D-peptides and D-proteins as potential therapeutic and informational tools.

DOI: 10.1038/s41557-023-01411-x

Source: https://www.nature.com/articles/s41557-023-01411-x