School of Chemistry and Chemical Engineering, South China University of Technology, Guangzhou 510640, China.
School of Chemistry and Chemical Engineering, South China University of Technology, Guangzhou 510640, China.
Bioorg Med Chem. 2024 Oct 1;112:117900. doi: 10.1016/j.bmc.2024.117900. Epub 2024 Aug 29.
The incorporation of non-canonical amino acids (ncAAs) into the metal coordination environments of proteins has endowed metalloproteins with enhanced properties and novel activities, particularly in hemoproteins. In this work, we disclose a scalable synthetic strategy that enables the production of myoglobin (Mb) variants with non-canonical heme ligands, i.e., HoCys and f4Tyr. The ncAA-containing Mb* variants (with H64V/V68A mutations) were obtained through two consecutive native chemical ligations and a subsequent desulfurization step, with overall isolated yield up to 28.6 % in over 10-milligram scales. After refolding and heme b cofactor reconstitution, the synthetic Mb* variants showed typical electronic absorption bands. When subjected to the catalysis of the cyclopropanation of styrene, both synthetic variants, however, were not as competent as the His-ligated Mb*. We envisioned that the synthetic method reported herein would be useful for incorporating a variety of ncAAs with diverse structures and properties into Mb for varied purposes.
将非天然氨基酸（ncAAs）整合到蛋白质的金属配位环境中，赋予了金属蛋白增强的性质和新的活性，特别是在血红素蛋白中。在这项工作中，我们揭示了一种可扩展的合成策略，能够生产具有非天然血红素配体的肌红蛋白（Mb）变体，即 HoCys 和 f4Tyr。含有 ncAA 的 Mb变体（具有 H64V/V68A 突变）通过两个连续的天然化学连接和随后的脱硫步骤获得，总体分离产率高达 28.6%，超过 10 毫克规模。在重折叠和血红素 b 辅因子重建后，合成的 Mb变体显示出典型的电子吸收带。然而，当进行苯乙烯的环丙烷化催化时，这两种合成变体都不如 His 连接的 Mb*有效。我们设想本文报道的合成方法将有助于将各种具有不同结构和性质的 ncAAs 整合到 Mb 中，用于不同的目的。
