Modularly engineering Rhodotorula toruloides for α-terpineol production

Liting Lyu; Qiongqiong Chen; Haizhao Xue; Sumayya Mustafa; Aabid Manzoor Shah; Qitian Huang; Yue Zhang; Shuang Wang; Zongbao Kent Zhao

doi:10.3389/fbioe.2023.1310069

Modularly engineering Rhodotorula toruloides for α-terpineol production

Front Bioeng Biotechnol. 2024 Jan 19:11:1310069. doi: 10.3389/fbioe.2023.1310069. eCollection 2023.

Authors

Liting Lyu^#^{1

2}, Qiongqiong Chen^#^{1

3}, Haizhao Xue^{1

3}, Sumayya Mustafa^{1

3}, Aabid Manzoor Shah¹, Qitian Huang¹, Yue Zhang^{1

3}, Shuang Wang^{1

3}, Zongbao Kent Zhao^{1

2}

Affiliations

¹ Laboratory of Biotechnology, Dalian Institute of Chemical Physics, Dalian, China.
² MOE Key Laboratory of Bio-Intelligent Manufacturing, School of Bioengineering, Dalian University of Technology, Dalian, China.
³ University of Chinese Academy of Sciences, Beijing, China.

^# Contributed equally.

Abstract

α-Terpineol is a monoterpenoid alcohol that has been widely used in the flavor, fragrance, and pharmaceutical industries because of its sensory and biological properties. However, few studies have focused on the microbial production of α-terpineol. The oleaginous yeast Rhodotorula toruloides is endowed with a natural mevalonate pathway and is a promising host in synthetic biology and biorefinery. The primary objective of this work was to engineer R. toruloides for the direct biosynthesis of α-terpineol. The improvement in monoterpenoid production was achieved through the implementation of modular engineering strategies, which included the enhancement of precursor supply, blocking of downstream pathways, and disruption of competing pathways. The results of these three methods showed varying degrees of favorable outcomes in enhancing α-terpineol production. The engineered strain 5L6HE5, with competitive pathway disruption and increased substrate supply, reached the highest product titer of 1.5 mg/L, indicating that reducing lipid accumulation is an efficient method in R. toruloides engineering for terpenoid synthesis. This study reveals the potential of R. toruloides as a host platform for the synthesis of α-terpineol as well as other monoterpenoid compounds.

Keywords: CRISPR/Cas9; Rhodotorula toruloides; modularly engineering; monoterpenoid; α-terpineol.

Grants and funding

The author(s) declare that financial support was received for the research, authorship, and/or publication of this article. This study was supported by the National Key Research and Development Program of China (2021YFA0910600), the National Natural Science Foundation of China (22308350 and 22238010), and the Liaoning Revitalization Talents Program (XLYC2002089).