Anal Chem. 2011 Feb 15;83(4):1202-6. doi: 10.1021/ac1032148. Epub 2011 Jan 27.
We report here the development of a straightforward, sensitive, and quantitative NMR-based method for high-throughput characterization of carbohydrate structure and screening of carbohydrate active enzyme (CAZyme) specificity. Automated assays starting from gene library expression to carbohydrate structure determination directly from crude reaction media have been established and successfully used to screen a library of 4032 CAZymes obtained by combinatorial engineering, at a rate of 480 enzyme variants per day. This allowed one to accurately discriminate 303 enzyme variants with altered specificity. The results demonstrate the potential of high-throughput NMR technology in glycomics, to mine artificial and natural enzyme diversity for novel biocatalysts.
我们在此报告了一种直接、灵敏、定量的基于 NMR 的方法的开发，用于高通量表征碳水化合物结构和筛选碳水化合物活性酶（CAZyme）特异性。从基因文库表达到直接从粗反应介质中确定碳水化合物结构的自动化测定已经建立，并成功用于筛选通过组合工程获得的 4032 种 CAZymes 文库，每天可筛选 480 种酶变体。这使得能够准确区分 303 种具有改变特异性的酶变体。结果表明，高通量 NMR 技术在糖组学中具有挖掘人工和天然酶多样性的潜力，用于寻找新型生物催化剂。
