Breeding of a Mesophilic Xanthophyllomyces dendrorhous Mutant for Industrial Astaxanthin Production and Mechanism Analysis on the Temperature Tolerance

Astaxanthin is a carotenoid with potent antioxidant properties, showing promising applications in cosmetics, animal feed, and human health. Xanthophyllomyces dendrorhous (also known as Phaffia rhodozyma) can indigenously synthesize astaxanthin with a broad substrate spectrum and robust growth, indicating significant potential for large-scale astaxanthin production. However, the low growth temperature (18–22 °C) and limited astaxanthin production levels hinder its industrial application. In this study, a combined mutagenesis and targeted screening strategy was established to successfully isolate mesophilic X. dendrorhous mutants capable of producing high levels of astaxanthin at 25 °C. Morphological, physiological, and biochemical investigations revealed that the enhanced membrane fluidity of the mutant strain contributed to its temperature tolerance. Further comparative genome and enzyme structure analyses identified mutations at bases 929 and 1017 of idh2 and bases 326 and 550 of fad3 as key factors for temperature tolerance, while mutations at base 130 of crtYB and base 406 of hmgr enhanced the astaxanthin synthesis. Additionally, transcriptional analysis revealed that genes related to the reactive oxygen species and NADPH modules were upregulated by 13-fold and 8.4-fold, respectively, promoting the astaxanthin synthesis in the mutant strain at 25 °C. Finally, mutant strain synthesized astaxanthin with a production of 1120 mg/L and content of 14 mg/g from sweet sorghum juice in 200 L fermenter. This represents the highest level of astaxanthin synthesis reported to date by X. dendrorhous, especially at 25 °C.