In vitro biosynthesis of poly-ß-1,4-glucan derivatives using a pro-miscuous glycosyltransferase
In vitro biosynthesis of poly-ß-1,4-glucan derivatives using a pro-miscuous glycosyltransferase
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使用混杂糖基转移酶体外生物合成聚-β-1,4-葡聚糖衍生物
DOI：
10.1101/2020.02.14.949545
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发表时间：
2020
期刊：
影响因子：
--
通讯作者：
Bulmer G
中科院分区：
文献类型：
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作者：
Bulmer G
纤维素的β-1，4-葡萄糖键是地球上最丰富的聚合键，因此在生物学和生物技术中具有相当大的兴趣。由于缺乏合适的生物催化剂，在体外合成这种连接仍然具有挑战性;天然纤维素生物合成机制是一种具有进行活性的膜相关复合物，不能容易地操纵以合成定制的寡糖及其衍生物。在这里，我们鉴定了可溶性重组生物催化剂脑膜炎奈瑟氏球菌二糖基转移酶LgtB的混杂活性，其适于通过产生β-1，4-糖苷键从UDP-葡萄糖聚合葡萄糖。我们采用LgtB合成天然和衍生的纤维寡糖，我们展示了如何LgtB可以被纳入生物催化级联和化学-酶促策略，以合成具有定制功能的纤维寡糖。我们还展示了如何得到的聚糖结构可以作为化学探针，以报告植物细胞壁降解酶，包括溶解性多糖单加氧酶的活性和选择性。我们预计，这种通过葡萄糖聚合衍生纤维低聚糖的生物催化方法将在生物学和纳米生物技术中开辟新的应用。
The β-1,4-glucose linkage of cellulose is the most abundant polymeric linkage on earth and as such is of considerable interest in biology and biotechnology. It remains challenging to synthesize this linkage in vitro due to a lack of suitable biocatalysts; the natural cellulose biosynthetic machinery is a membrane-associated complex with processive activity that cannot be easily manipulated to synthesize tailor-made oligosaccharides and their derivatives. Here we identify a promiscuous activity of a soluble recombinant biocatalyst,Neisseria meningitidisglycosyltransferase LgtB, suitable for the polymerization of glucose from UDP-glucose via the generation of β-1,4-glycosidic linkages. We employed LgtB to synthesize natural and derivatized cello-oligosaccharides and we demonstrate how LgtB can be incorporated in biocatalytic cascades and chemo-enzymatic strategies to synthesize cello-oligosaccharides with tailored functionalities. We also show how the resulting glycan structures can be applied as chemical probes to report on activity and selectivity of plant cell wall degrading enzymes, including lytic polysaccharide monooxygenases. We anticipate that this biocatalytic approach to derivatized cello-oligosaccharides via glucose polymerization will open up new applications in biology and nanobiotechnology.