Biocatalytic Transfer of Pseudaminic Acid (Pse5Ac7Ac) Using Promiscuous Sialyltransferases in a Chemoenzymatic Approach to Pse5Ac7Ac-Containing Glycosides
Biocatalytic Transfer of Pseudaminic Acid (Pse5Ac7Ac) Using Promiscuous Sialyltransferases in a Chemoenzymatic Approach to Pse5Ac7Ac-Containing Glycosides
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DOI：
10.1021/acscatal.0c02189
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发表时间：
2020-09-04
期刊：
影响因子：
12.9
通讯作者：
Fascione, Martin A.
中科院分区：
文献类型：
--
作者：
Flack, Emily K. P.;Chidwick, Harriet S.;Fascione, Martin A.
假氨基酸（Pse 5Ac 7Ac）是一种存在于许多细菌细胞表面的非哺乳动物糖，包括铜绿假单胞菌、空肠弯曲杆菌和鲍曼不动杆菌。然而，Pse 5Ac 7Ac在宿主-病原体相互作用中所起的作用仍然未被探索，特别是与其普遍存在的唾液酸类似物Neu 5Ac相比。这主要是由于缺乏获得难以制备的Pse 5Ac 7Ac糖苷的途径。在此，我们描述了体外生物催化转移的活化Pse 5Ac 7Ac供体糖基受体，使酶促合成的Pse 5Ac 7Ac-含有糖苷。在化学酶促方法中，化学合成最初提供了后期Pse 5Ac 7Ac生物合成中间体的途径，随后使用生物合成酶在一锅双酶过程中将其转化为所需的CMP-糖基供体。最后，用非天然底物筛选13种唾液酸转移酶（SiaT）的文库，使得能够鉴定能够周转以提供β-Pse 5Ac 7Ac-终止的糖苷的混杂的倒位SiaT。
Pseudaminic acid (Pse5Ac7Ac) is a nonmammalian sugar present on the cell surface of a number of bacteria including Pseudomonas aeruginosa, Campylobacter jejuni, and Acinetobacter baumannii. However, the role Pse5Ac7Ac plays in host-pathogen interactions remains underexplored, particularly compared to its ubiquitous sialic acid analogue Neu5Ac. This is primarily due to a lack of access to difficult to prepare Pse5Ac7Ac glycosides. Herein, we describe the in vitro biocatalytic transfer of an activated Pse5Ac7Ac donor onto glycosyl acceptors, enabling the enzymatic synthesis of Pse5Ac7Ac-containing glycosides. In a chemoenzymatic approach, chemical synthesis initially afforded access to a late-stage Pse5Ac7Ac biosynthetic intermediate, which was subsequently converted to the desired CMP-glycosyl donor in a one-pot two-enzyme process using biosynthetic enzymes. Finally, screening a library of 13 sialyltransferases (SiaT) with the unnatural substrate enabled the identification of a promiscuous inverting SiaT capable of turnover to afford beta-Pse5Ac7Ac-terminated glycosides.