TY - JOUR
T1 - Cas9-Based Metabolic Engineering of Issatchenkia orientalis for Enhanced Utilization of Cellulosic Hydrolysates
AU - Lee, Ye Gi
AU - Kim, Chanwoo
AU - Kuanyshev, Nurzhan
AU - Kang, Nam Kyu
AU - Fatma, Zia
AU - Wu, Zong Yen
AU - Cheng, Ming Hsun
AU - Singh, Vijay
AU - Yoshikuni, Yasuo
AU - Zhao, Huimin
AU - Jin, Yong Su
N1 - Funding Information:
This work was supported by the DOE Center for Advanced Bioenergy and Bioproducts Innovation (U.S. Department of Energy, Office of Science, Office of Biological and Environmental Research under Award Number DE-SC0018420 and DE-AC02-05CH11231). The work conducted by the U.S. Department of Energy Joint Genome Institute, a DOE Office of Science User Facility, was supported under Contract No. DE-AC02-05CH11231. Any opinions, findings, and conclusions, or recommendations expressed in this publication are those of the author(s) and do not necessarily reflect the views of the U.S. Department of Energy.
Publisher Copyright:
© 2022 American Chemical Society.
PY - 2022/9/28
Y1 - 2022/9/28
N2 - Issatchenkia orientalis, exhibiting high tolerance against harsh environmental conditions, is a promising metabolic engineering host for producing fuels and chemicals from cellulosic hydrolysates containing fermentation inhibitors under acidic conditions. Although genetic tools for I. orientalis exist, they require auxotrophic mutants so that the selection of a host strain is limited. We developed a drug resistance gene (cloNAT)-based genome-editing method for engineering any I. orientalis strains and engineered I. orientalis strains isolated from various sources for xylose fermentation. Specifically, xylose reductase, xylitol dehydrogenase, and xylulokinase from Scheffersomyces stipitis were integrated into an intended chromosomal locus in four I. orientalis strains (SD108, IO21, IO45, and IO46) through Cas9-based genome editing. The resulting strains (SD108X, IO21X, IO45X, and IO46X) efficiently produced ethanol from cellulosic and hemicellulosic hydrolysates even though the pH adjustment and nitrogen source were not provided. As they presented different fermenting capacities, selection of a host I. orientalis strain was crucial for producing fuels and chemicals using cellulosic hydrolysates.
AB - Issatchenkia orientalis, exhibiting high tolerance against harsh environmental conditions, is a promising metabolic engineering host for producing fuels and chemicals from cellulosic hydrolysates containing fermentation inhibitors under acidic conditions. Although genetic tools for I. orientalis exist, they require auxotrophic mutants so that the selection of a host strain is limited. We developed a drug resistance gene (cloNAT)-based genome-editing method for engineering any I. orientalis strains and engineered I. orientalis strains isolated from various sources for xylose fermentation. Specifically, xylose reductase, xylitol dehydrogenase, and xylulokinase from Scheffersomyces stipitis were integrated into an intended chromosomal locus in four I. orientalis strains (SD108, IO21, IO45, and IO46) through Cas9-based genome editing. The resulting strains (SD108X, IO21X, IO45X, and IO46X) efficiently produced ethanol from cellulosic and hemicellulosic hydrolysates even though the pH adjustment and nitrogen source were not provided. As they presented different fermenting capacities, selection of a host I. orientalis strain was crucial for producing fuels and chemicals using cellulosic hydrolysates.
KW - Cas9 genome-editing strategies
KW - Issatchenkia orientalis
KW - bioenergy sorghum
KW - no pH adjustment
KW - nonconventional yeast
KW - xylose fermentation
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U2 - 10.1021/acs.jafc.2c04251
DO - 10.1021/acs.jafc.2c04251
M3 - Article
C2 - 36103687
AN - SCOPUS:85138859311
SN - 0021-8561
VL - 70
SP - 12085
EP - 12094
JO - Journal of Agricultural and Food Chemistry
JF - Journal of Agricultural and Food Chemistry
IS - 38
ER -