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Major aldehyde dehydrogenase AldFGH of Gluconacetobacter diazotrophicus is independent of pyrroloquinoline quinone but dependent on molybdopterin for acetic acid fermentation

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Abstract

Acetic acid fermentation involves the oxidation of ethanol to acetic acid via acetaldehyde as the intermediate and is catalyzed by the membrane-bound alcohol dehydrogenase (ADH) and aldehyde dehydrogenase (ALDH) of acetic acid bacteria. Although ADH depends on pyrroloquinoline quinone (PQQ), the prosthetic group associated with ALDH remains a matter of debate. This study aimed to address the dependency of ALDH of Gluconacetobacter diazotrophicus strain PAL5 on PQQ and the physiological role of ALDH in acetic acid fermentation. We constructed deletion mutant strains for both the ALDH gene clusters of PAL5, aldFGH and aldSLC. In addition, the adhAB operon for ADH was eliminated, since it shows ALDH activity. The triple-deletion derivative ΔaldFGH ΔaldSLC ΔadhAB failed to show ALDH activity, which suggested that ALDH activity in PAL5 is derived from these three enzyme complexes. Since the single-gene cluster deletion derivative ΔaldFGH lost most ALDH activity, and accumulated much higher acetaldehyde than wild type under acetic acid fermentation conditions, we concluded that AldFGH functions as the major ALDH in PAL5. Furthermore, deletion of the PQQ biosynthesis gene cluster (pqqABCDE) abolished ADH activity completely, but did not affect ALDH activity. Instead, the molybdopterin biosynthesis gene deletion derivatives lost ALDH activity. Thus, we concluded that the AldFGH and AldSLC complexes of Ga. diazotrophicus PAL5 require a form of molybdopterin but not PQQ for ALDH activity.

Key points

• AldFGH is the major aldehyde dehydrogenase in Gluconacetobacter diazotrophicus PAL5.

• Acetaldehyde accumulated from ethanol in the absence of AldFGH.

• Molybdopterin, rather than pyrroloquinoline quinone, is required for AldFGH.

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Data availability

The data that support the findings of this study are available from the corresponding author upon reasonable request.

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Acknowledgments

We are grateful to the Japanese Government (Monbukagakusho: MEXT) Scholarships that supported RM work in this study. We thank Oriental Yeast (Tokyo, Japan) and Toyobo (Osaka, Japan) for gifting us with yeast extract and restriction endonucleases, respectively. We are grateful to Tomoyuki Kosaka and Mamoru Yamada for their invaluable suggestions. We thank Osao Adachi and Hirohide Toyama for encouraging our ALDH work. This work was supported by KAKENHI grant number 23580115.

Funding

This work was supported by KAKENHI grant number 23580115.

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Authors and Affiliations

  1. Graduate School of Sciences and Technology for Innovation, Yamaguchi University, Yamaguchi, 753-8515, Japan

    Roni Miah, Takeru Murate, Naoya Kataoka, Kazunobu Matsushita & Toshiharu Yakushi

  2. Faculty of Agriculture, Yamaguchi University, Yamaguchi, 753-8515, Japan

    Shun Nina, Naoya Kataoka, Kazunobu Matsushita & Toshiharu Yakushi

  3. Research Center for Thermotolerant Microbial Resources, Yamaguchi University, Yamaguchi, 753-8515, Japan

    Naoya Kataoka, Kazunobu Matsushita & Toshiharu Yakushi

  4. NODAI Genome Research Center, Tokyo University of Agriculture, Tokyo, 156-8502, Japan

    Minenosuke Matsutani

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  1. Roni Miah
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  2. Shun Nina
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Contributions

TY designed the study and wrote the manuscript. RM performed most experiments, analyzed the data, prepared the figures, and wrote a draft manuscript. SN and TM assisted in constructing bacterial strains and editing the manuscript. MM performed bioinformatics analysis. TY, NK, and KM supervised and edited the manuscript. All authors have read and approved the manuscript.

Corresponding author

Correspondence to Toshiharu Yakushi.

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Miah, R., Nina, S., Murate, T. et al. Major aldehyde dehydrogenase AldFGH of Gluconacetobacter diazotrophicus is independent of pyrroloquinoline quinone but dependent on molybdopterin for acetic acid fermentation. Appl Microbiol Biotechnol 105, 2341–2350 (2021). https://doi.org/10.1007/s00253-021-11144-x

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