Davenport Chemistry Laboratories, Department of Chemistry, University of Toronto, Toronto, Ontario M5S 3H6, Canada.
Org Biomol Chem. 2022 Oct 26;20(41):8083-8091. doi: 10.1039/d2ob01372j.
Chemical cross-linking of human adult hemoglobin (Hb) prevents dissociation of the tetrameric (αβ) protein into its constituent non-functional αβ dimers when present outside red cells, providing the possibility of being an acellular oxygen carrier in circulation. However, studies of cross-linked Hb (xlHb) in circulation established effects consistent with scavenging of endogenous nitric oxide, leading to hypertension. Bis-tetramers, composed of coupled Hb tetramers, are sufficiently large to avoid penetration of endothelia, thereby blocking access to endogenous nitric oxide. Cu(I)-catalyzed azide-alkyne cycloaddition (CuAAC) joins two azide-functionalized xlHbs to each end of a bis-alkyne to form bis-tetramers. The process critically depends on formation of a cross-link between lysyl amino groups of the β-subunits while avoiding reactions with amino groups in the α-subunits. Highly selective acetylation of α-subunit amino groups with methyl acetyl phosphate (MAP) effectively directs subsequent cross-linking to the β-subunits. This outcome leads to efficient production of hemoglobin bis-tetramers by CuAAC.
人成年血红蛋白（Hb）的化学交联可防止四聚体（αβ）蛋白在红细胞外解聚成无功能的αβ二聚体，从而有可能成为循环中的无细胞氧载体。然而，对循环中交联血红蛋白（xlHb）的研究表明，其具有清除内源性一氧化氮的作用，导致高血压。双四聚体由偶联的 Hb 四聚体组成，其足够大，可避免穿透内皮细胞，从而阻止内源性一氧化氮进入。Cu（I）催化的叠氮-炔环加成（CuAAC）将两个叠氮功能化的 xlHb 连接到双炔的两端，形成双四聚体。该过程严重依赖于β亚基赖氨酸氨基之间形成交联，同时避免与α亚基中的氨基反应。用甲基乙酰膦酸盐（MAP）高度选择性乙酰化α-亚基氨基可有效地将随后的交联导向β-亚基。这一结果导致通过 CuAAC 有效地生产血红蛋白双四聚体。
