TY - JOUR
T1 - Metabolic engineering of low-pH-tolerant non-model yeast, Issatchenkia orientalis, for production of citramalate
AU - Wu, Zong Yen
AU - Sun, Wan
AU - Shen, Yihui
AU - Pratas, Jimmy
AU - Suthers, Patrick F.
AU - Hsieh, Ping Hung
AU - Dwaraknath, Sudharsan
AU - Rabinowitz, Joshua D.
AU - Maranas, Costas D.
AU - Shao, Zengyi
AU - Yoshikuni, Yasuo
N1 - Funding Information:
This work was funded 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 ( https://ror.org/04xm1d337 ), a DOE Office of Science User Facility, is supported by the Office of Science of the U.S. Department of Energy operated 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. We thank Anita Wahler for professional editing.
Publisher Copyright:
© 2023 The Authors
PY - 2023/6
Y1 - 2023/6
N2 - Methyl methacrylate (MMA) is an important petrochemical with many applications. However, its manufacture has a large environmental footprint. Combined biological and chemical synthesis (semisynthesis) may be a promising alternative to reduce both cost and environmental impact, but strains that can produce the MMA precursor (citramalate) at low pH are required. A non-conventional yeast, Issatchenkia orientalis, may prove ideal, as it can survive extremely low pH. Here, we demonstrate the engineering of I. orientalis for citramalate production. Using sequence similarity network analysis and subsequent DNA synthesis, we selected a more active citramalate synthase gene (cimA) variant for expression in I. orientalis. We then adapted a piggyBac transposon system for I. orientalis that allowed us to simultaneously explore the effects of different cimA gene copy numbers and integration locations. A batch fermentation showed the genome-integrated-cimA strains produced 2.0 g/L citramalate in 48 h and a yield of up to 7% mol citramalate/mol consumed glucose. These results demonstrate the potential of I. orientalis as a chassis for citramalate production.
AB - Methyl methacrylate (MMA) is an important petrochemical with many applications. However, its manufacture has a large environmental footprint. Combined biological and chemical synthesis (semisynthesis) may be a promising alternative to reduce both cost and environmental impact, but strains that can produce the MMA precursor (citramalate) at low pH are required. A non-conventional yeast, Issatchenkia orientalis, may prove ideal, as it can survive extremely low pH. Here, we demonstrate the engineering of I. orientalis for citramalate production. Using sequence similarity network analysis and subsequent DNA synthesis, we selected a more active citramalate synthase gene (cimA) variant for expression in I. orientalis. We then adapted a piggyBac transposon system for I. orientalis that allowed us to simultaneously explore the effects of different cimA gene copy numbers and integration locations. A batch fermentation showed the genome-integrated-cimA strains produced 2.0 g/L citramalate in 48 h and a yield of up to 7% mol citramalate/mol consumed glucose. These results demonstrate the potential of I. orientalis as a chassis for citramalate production.
UR - http://www.scopus.com/inward/record.url?scp=85148677011&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=85148677011&partnerID=8YFLogxK
U2 - 10.1016/j.mec.2023.e00220
DO - 10.1016/j.mec.2023.e00220
M3 - Article
C2 - 36860699
AN - SCOPUS:85148677011
SN - 2214-0301
VL - 16
JO - Metabolic Engineering Communications
JF - Metabolic Engineering Communications
M1 - e00220
ER -
