LAUSR

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.