Tao Xuanyu, Morgan Josiah S, Liu Jiantao, Kempher Megan L, Xu Tao, Zhou Jizhong
Institute for Environmental Genomics, Department of Microbiology and Plant Biology, and School of Civil Engineering and Environmental Sciences, University of Oklahoma, Norman, OK, USA.
Institute for Environmental Genomics, Department of Microbiology and Plant Biology, and School of Civil Engineering and Environmental Sciences, University of Oklahoma, Norman, OK, USA; Earth and Environmental Sciences, Lawrence Berkeley National Laboratory, Berkeley, CA, USA.
Bioresour Technol. 2023 May;376:128849. doi: 10.1016/j.biortech.2023.128849. Epub 2023 Mar 9.
The bacteria Clostridium cellulolyticum is a promising candidate for consolidated bioprocessing (CBP). However, genetic engineering is necessary to improve this organism's cellulose degradation and bioconversion efficiencies to meet standard industrial requirements. In this study, CRISPR-Cas9n was used to integrate an efficient β-glucosidase into the genome of C. cellulolyticum, disrupting lactate dehydrogenase (ldh) expression and reducing lactate production. The engineered strain showed a 7.4-fold increase in β-glucosidase activity, a 70% decrease in ldh expression, a 12% increase in cellulose degradation, and a 32% increase in ethanol production compared to wild type. Additionally, ldh was identified as a potential site for heterologous expression. These results demonstrate that simultaneous β-glucosidase integration and lactate dehydrogenase disruption is an effective strategy for increasing cellulose to ethanol bioconversion rates in C. cellulolyticum.
解纤维素梭菌是用于整合生物加工（CBP）的一个很有前景的候选菌株。然而，有必要通过基因工程来提高该生物体的纤维素降解和生物转化效率，以满足标准的工业要求。在本研究中，使用CRISPR-Cas9n将一种高效的β-葡萄糖苷酶整合到解纤维素梭菌的基因组中，破坏乳酸脱氢酶（ldh）的表达并减少乳酸生成。与野生型相比，工程菌株的β-葡萄糖苷酶活性提高了7.4倍，ldh表达降低了70%，纤维素降解增加了12%，乙醇产量增加了32%。此外，ldh被确定为异源表达的一个潜在位点。这些结果表明，同时整合β-葡萄糖苷酶和破坏乳酸脱氢酶是提高解纤维素梭菌中纤维素到乙醇生物转化率的有效策略。
