Biocatalysis of heterogenously-expressed d-lactonohydrolases and its efficient preparation of desirable d-pantoic acid

• Published in: Enzyme and microbial technology Vol. 155; p. 109981
• Main Authors: Sun, Ruobin, Zheng, Pu, Wu, Dan, Chen, Pengcheng, Bai, Yanbing, Wang, Jun
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 01.04.2022
• Subjects: Biocatalysis; D-Lactonohydrolase; D-Pantoic acid; Enzymatic resolution; Heterologous expression; Hydrolysis; Hydroxybutyrates; Pichia - genetics; Pichia - metabolism; Pichia pastoris; Recombinant Proteins - genetics; Recombinant Proteins - metabolism; D-Lactonohydrolase; Enzymatic resolution; D-Pantoic acid; Pichia pastoris; Heterologous expression
• ISSN: 0141-0229; 1879-0909
• DOI: 10.1016/j.enzmictec.2021.109981

d-Pantoic acid (D-PA) is an essential intermediate for the production of d-pantolactone. Here, three d-lactonohydrolases (D-Lacs), namely, Fm-Lac from Fusarium moniliforme SW-902, Fp-Lac from Fusarium proliferatum Nirenberg ECU2002, and Fo-Lac from Fusarium oxysporum AKU3702 were heterogeneously expressed in Pichia pastoris. The constructed recombinant strains produced D-Lacs of 1263 U/mL, 1025 U/mL, and 948 U/mL in a 3-L fermenter, respectively. Simultaneously, these three D-Lacs were used to resolve racemic pantolactone (DL-PL), the hydrolysis rate by Fo-Lac over 40% and the enantiomeric excesses was 99% after 4 h reaction, which outperformed Fm-Lac and Fp-Lac. Under the 800 mL scale reaction, the hydrolysis rate of DL-PL reached 39.2% with a D-PA concentration of 144.6 g/L and space-time yield of 36.2 g/L/h correspondingly. This is the highest catalytic efficiency reported so far, which shows that D-Lac heterologously expressed by P. pastoris has excellent industrial application prospects. •D-Lac was successfully expressed in Pichia pastoris for the first time.•The production of D-Lac by the recombinant P. pastoris is as high as 1000 U/mL or more.•The space-time yield of D-PA production reached 36.2 g/L/h, which is currently the highest in the resolution of DL-PL.