A biosynthetic pathway for hexanoic acid production in Kluyveromyces marxianus.
Hexanoic acid can be used for diverse industrial applications and is a precursor for fine chemistry. Although some natural microorganisms have been screened and evolved to produce hexanoic acid, the construction of an engineered biosynthetic pathway for producing hexanoic acid in yeast has not been reported. Here we constructed hexanoic acid pathways in Kluyveromyces marxianus by integrating 5 combinations of seven genes (AtoB, BktB, Crt, Hbd, MCT1, Ter, and TES1), by which random chromosomal sites of the strain are overwritten by the new genes from bacteria and yeast. One recombinant strain, H4A, which contained AtoB, BktB, Crt, Hbd, and Ter, produced 154mg/L of hexanoic acid from galactose as the sole substrate. However, the hexanoic acid produced by the H4A strain was re-assimilated during the fermentation due to the reverse activity of AtoB, which condenses two acetyl-CoAs into a single acetoacetyl-CoA. This product instability could be overcome by the replacement of AtoB with a malonyl CoA-acyl carrier protein transacylase (MCT1) from Saccharomyces cerevisiae. Our results suggest that Mct1 provides a slow but stable acetyl-CoA chain elongation pathway, whereas the AtoB-mediated route is fast but unstable. In conclusion, hexanoic acid was produced for the first time in yeast by the construction of chain elongation pathways comprising 5-7 genes in K. marxianus.
Department of Genetic Engineering, College of Biotechnology and Bioengineering, Sungkyunkwan University, Suwon 440-746, Republic of Korea.
Yuna
Y
Cheon
Department of Genetic Engineering, College of Biotechnology and Bioengineering, Sungkyunkwan University, Suwon 440-746, Republic of Korea.
Jun-Seob
JS
Kim
Department of Genetic Engineering, College of Biotechnology and Bioengineering, Sungkyunkwan University, Suwon 440-746, Republic of Korea.
Jun-Bum
JB
Park
Department of Genetic Engineering, College of Biotechnology and Bioengineering, Sungkyunkwan University, Suwon 440-746, Republic of Korea.
Paul
P
Heo
School of Interdisciplinary Bioscience and Bioengineering, Pohang University of Science and Technology, Pohang 790-784, Republic of Korea.
Jae Hyung
JH
Lim
School of Interdisciplinary Bioscience and Bioengineering, Pohang University of Science and Technology, Pohang 790-784, Republic of Korea; Department of Chemical Engineering, Pohang University of Science and Technology, Pohang 790-784, Republic of Korea.
Gyoo Yeol
GY
Jung
Department of Agricultural Biotechnology and Center for Agricultural Biomaterials, Seoul National University, Seoul 151-921, Republic of Korea.
Jin-Ho
JH
Seo
Bio R&D Center, Samsung Advanced Institute of Technology, Yongin 446-712, Republic of Korea.
Jin Hwan
JH
Park
Bio R&D Center, Samsung Advanced Institute of Technology, Yongin 446-712, Republic of Korea.
Hyun Min
HM
Koo
Bio R&D Center, Samsung Advanced Institute of Technology, Yongin 446-712, Republic of Korea.
Kwang Myung
KM
Cho
Department of Food Science and Engineering, Ewha Womans University, Seoul 120-750, Republic of Korea.
Jin-Byung
JB
Park
Department of Bioengineering and Technology, College of Engineering, Kangwon National University, Chuncheon 220-701, Republic of Korea. Electronic address: sjha@kangwon.ac.kr.
Suk-Jin
SJ
Ha
Department of Genetic Engineering, College of Biotechnology and Bioengineering, Sungkyunkwan University, Suwon 440-746, Republic of Korea. Electronic address: dhkweon@skku.edu.
Dae-Hyuk
DH
Kweon
D001709:Biotechnology; D002208:Caproates / Q000032:analysis / Q000378:metabolism*; D000431:Ethanol / Q000378:metabolism; D005285:Fermentation; D005690:Galactose / Q000378:metabolism; D005947:Glucose / Q000378:metabolism; D007716:Kluyveromyces / Q000235:genetics* / Q000378:metabolism*; D060847:Metabolic Engineering / Q000379:methods*; D053858:Metabolic Networks and Pathways
D002208:Caproates; C037652:hexanoic acid; D000431:Ethanol; D005947:Glucose; D005690:Galactose
AtoB; Hexanoic acid; Kluyveromyces marxianus; MCT1; TES1