Department of Energy Great Lakes Bioenergy Research Center and Department of Energy Plant Research Laboratory, Michigan State University, E. Lansing, MI 48824, USA.
Bioresour Technol. 2010 Dec;101(23):9097-105. doi: 10.1016/j.biortech.2010.07.028. Epub 2010 Jul 13.
A high throughput enzyme assay platform, called GENPLAT, was used to guide the development of an optimized mixture of individual purified enzymes from ten "accessory" and six "core" enzymes. Enzyme mixtures were optimized for release of Glu, Xyl, or a combination of the two from corn stover pretreated by ammonia-fiber expansion (AFEX). Assay conditions were a fixed enzyme loading of 15 mg/g glucan, 48 h digestion, and 50 degrees C. Five of the ten tested accessory proteins enhanced Glu or Xyl yield compared to the core set alone, and five did not. An 11-component mixture containing the core set and five accessory enzymes optimized for Glu released 52.1% of the available Glu, compared to 38.5% with the core set alone. A mixture optimized for Xyl released 39.9% of the Xyl, compared to 26.4% with the core set alone. We predict that there is still considerable opportunity for further improvement of synthetic mixtures. Furthermore, the strategy described here is applicable to the development of more efficient enzyme cocktails for any pretreatment/biomass combination and for detecting enzymes that make a heretofore unrecognized contribution to lignocellulose deconstruction.
高通量酶分析平台 GENPLAT 用于指导从十种“辅助”和六种“核心”酶中优化组合单独的纯化酶。酶混合物优化了从氨纤维膨胀预处理的玉米秸秆中释放 Glu、Xyl 或两者的混合物。分析条件为固定酶负载量为 15mg/g 葡聚糖，48 小时消化，50°C。在测试的十种辅助蛋白中，有五种与核心组相比提高了 Glu 或 Xyl 的产量，而有五种没有。含有核心组和五个辅助酶的 11 种酶混合物优化 Glu 释放，释放了 52.1%的可用 Glu，而核心组单独释放了 38.5%。优化 Xyl 的混合物释放了 39.9%的 Xyl，而核心组单独释放了 26.4%。我们预测，进一步提高合成混合物的效率仍有很大的机会。此外，这里描述的策略适用于开发任何预处理/生物质组合的更有效的酶混合物，以及检测到迄今为止对木质纤维素解构做出未被认识到的贡献的酶。
