Kim Youngmi, Kreke Thomas, Ko Ja Kyong, Ladisch Michael R
Laboratory of Renewable Resources Engineering, Purdue University, West Lafayette, Indiana; Department of Agricultural and Biological Engineering, Purdue University, West Lafayette, Indiana.
Biotechnol Bioeng. 2015 Apr;112(4):677-87. doi: 10.1002/bit.25465. Epub 2015 Jan 2.
Fundamental characterization of pretreated hardwood and its interactions with cellulolytic enzymes has confirmed that a pathway exists for dramatically reducing the loading of cellulase required for hydrolysis of pretreated biomass. We demonstrate that addition of protein effecting a seven-fold decrease in the specific activity of cellulases enables a ten-fold reduction in enzyme loading while maintaining a high level of cellulose hydrolysis in pretreated hardwood. While use of protein and other additives that adsorb on lignin have been reported previously, the current work demonstrates the effect in a dramatic manner and brings the rationale for this change into clear focus. The key to this result is recognizing and mitigating the pretreatment conundrum where increasingly severe pretreatment conditions enhance accessibility of the enzymes not only to cellulose, but also to lignin. The lignin adsorbs enzyme protein causing loss of cellulase activity. More enzyme, added to compensate for this lost activity, results in a higher cellulase loading. The addition of a different protein, such as BSA, prevents cellulase adsorption on lignin and enables the enzyme itself to better target its glucan substrate. This effect dramatically reduces the amount of cellulase for a given level of conversion with enzyme loadings of 15 FPU and 1.3 FPU/g solids both achieving 80% conversion. The understanding of this phenomenon reinvigorates motivation for the search for other approaches that prevent cellulase adsorption on lignin in order to achieve high glucose yields at low enzyme loadings for pretreated lignocellulose.
预处理硬木的基本特性及其与纤维素分解酶的相互作用已证实，存在一条途径可显著降低预处理生物质水解所需纤维素酶的用量。我们证明，添加能使纤维素酶比活性降低七倍的蛋白质，可使酶用量减少十倍，同时在预处理硬木中保持高水平的纤维素水解。虽然此前已有报道使用吸附在木质素上的蛋白质和其他添加剂，但当前的研究以显著的方式展示了这种效果，并使这种变化的原理清晰地凸显出来。这一结果的关键在于认识并缓解预处理难题，即日益严格的预处理条件不仅提高了酶对纤维素的可及性，也提高了对木质素的可及性。木质素吸附酶蛋白会导致纤维素酶活性丧失。为补偿这种活性损失而添加更多的酶，会导致更高的纤维素酶用量。添加不同的蛋白质，如牛血清白蛋白（BSA），可防止纤维素酶吸附在木质素上，并使酶本身能更好地靶向其葡聚糖底物。这种效果在给定的转化水平下显著减少了纤维素酶的用量，酶用量为15 FPU和1.3 FPU/g固体时均能实现80%的转化率。对这一现象的理解重新激发了人们寻找其他方法的动力，这些方法可防止纤维素酶吸附在木质素上，从而在低酶用量下实现预处理木质纤维素的高葡萄糖产率。
