Heterologous expression of membrane-bound alcohol dehydrogenase–encoding genes for glyceric acid production using Gluconobacter sp. CHM43 and its derivatives.
In: Applied Microbiology & Biotechnology, Jg. 105 (2021-09-01), Heft 18, S. 6749-6758
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Zugriff:
In contrast to D-glyceric acid (D-GA) production with 99% enantiomeric excess (ee) by Acetobacter tropicalis NBRC 16470, Gluconobacter sp. CHM43 produced 19.6 g L−1 of D-GA with 73.7% ee over 4 days of incubation in flask culture. To investigate the reason for this enantiomeric composition of GA, the genes encoding membrane-bound alcohol dehydrogenase (mADH) of A. tropicalis NBRC 16470, composed of three subunits (adhA, adhB, and adhS), were cloned using the broad-host-range vector pBBR1MCS-2 and heterologously expressed in Gluconobacter sp. CHM43 and its ΔadhAB ΔsldBA derivative TORI4. Reverse-transcription quantitative real-time polymerase chain reaction demonstrated that adhABS genes from A. tropicalis were expressed in TORI4 transformants, and their membrane fraction exhibited mADH activities of 0.13 and 0.31 U/mg with or without AdhS, respectively. Compared with the GA production of TORI4-harboring pBBR1MCS-2 (1.23 g L−1), TORI4 transformants expressing adhABS and adhAB showed elevated GA production of 2.46 and 3.67 g L−1, respectively, suggesting a negative effect of adhS gene expression on GA production as well as mADH activity in TORI4. Although TORI4 was found to produce primarily L-GA with 42.5% ee, TORI4 transformants expressing adhABS and adhAB produced D-GA with 27.6% and 49.0% ee, respectively, demonstrating that mADH of A. tropicalis causes a sharp increase in the enantiomeric composition of D-GA. These results suggest that one reason for D-GA production with 73.7% ee in Gluconobacter spp. might be a property of the host, which possibly produces L-GA intracellularly. Key points: • Membrane-bound ADH from Acetobacter tropicalis showed activity in Gluconobacter sp. • D-GA production from glycerol was performed using recombinant Gluconobacter sp. • Enantiomeric excess of D-GA was affected by both membrane and intracellular ADHs. [ABSTRACT FROM AUTHOR]
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Heterologous expression of membrane-bound alcohol dehydrogenase–encoding genes for glyceric acid production using Gluconobacter sp. CHM43 and its derivatives.
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Autor/in / Beteiligte Person: | Habe, Hiroshi ; Sato, Yuya ; Tani, Hidenori ; Matsutani, Minenosuke ; Tanioka, Kazuya ; Theeragool, Gunjana ; Matsushita, Kazunobu ; Yakushi, Toshiharu |
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Zeitschrift: | Applied Microbiology & Biotechnology, Jg. 105 (2021-09-01), Heft 18, S. 6749-6758 |
Veröffentlichung: | 2021 |
Medientyp: | academicJournal |
ISSN: | 0175-7598 (print) |
DOI: | 10.1007/s00253-021-11535-0 |
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