한빛사 논문
Dae-yeol Ye1,4, Myung Hyun Noh1,4, Jo Hyun Moon1,4, Alfonsina Milito2, Minsun Kim3, Jeong Wook Lee1,3, Jae-Seong Yang2,* & Gyoo Yeol Jung1,3,*
1Department of Chemical Engineering, Pohang University of Science and Technology, 77 Cheongam-Ro, Nam-Gu, Pohang, Gyeongbuk 37673, Republic of Korea. 2Centre for Research in Agricultural Genomics (CRAG), CSIC-IRTA-UAB-UB, Campus UAB, Bellaterra, Barcelona 08193, Spain. 3School of Interdisciplinary Bioscience and Bioengineering, Pohang University of Science and Technology, 77 Cheongam-Ro, Nam-Gu, Pohang, Gyeongbuk 37673, Republic of Korea. 4These authors contributed equally: Dae-yeol Ye, Myung Hyun Noh, Jo Hyun Moon.
*Corresponding author.
Abstract
Physical compartmentalization of metabolism using membranous organelles in eukaryotes is helpful for chemical biosynthesis to ensure the availability of substrates from competitive metabolic reactions. Bacterial hosts lack such a membranous system, which is one of the major limitations for efficient metabolic engineering. Here, we employ kinetic compartmentalization with the introduction of an unnatural enzymatic reaction by an engineered enzyme as an alternative strategy to enable substrate availability from competitive reactions through kinetic isolation of metabolic pathways. As a proof of concept, we kinetically isolate the itaconate synthetic pathway from the tricarboxylic acid cycle in Escherichia coli, which is natively separated by mitochondrial membranes in Aspergillus terreus. Specifically, 2-methylcitrate dehydratase is engineered to alternatively catalyze citrate and kinetically secure cis-aconitate for efficient production using a high-throughput screening system. Itaconate production can be significantly improved with kinetic compartmentalization and its strategy has the potential to be widely applicable.
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