Abstract
Lactobacillus reuteri, a heterofermentative bacterium, metabolizes glycerol via a Pdu (propanediol-utilization) pathway involving dehydration to 3-hydroxypropionaldehyde (3-HPA) followed by reduction to 1,3-propandiol (1,3-PDO) with concomitant generation of an oxidized cofactor, NAD+ that is utilized to maintain cofactor balance required for glucose metabolism and even for oxidation of 3-HPA by a Pdu oxidative branch to 3-hydroxypropionic acid (3-HP). The Pdu pathway is operative inside Pdu microcompartment that encapsulates different enzymes and cofactors involved in metabolizing glycerol or 1,2-propanediol, and protects the cells from the toxic effect of the aldehyde intermediate. Since L. reuteri excretes high amounts of 3-HPA outside the microcompartment, the organism is likely to have alternative alcohol dehydrogenase(s) in the cytoplasm for transformation of the aldehyde. In this study, diversity of alcohol dehydrogenases in Lactobacillus species was investigated with a focus on L. reuteri. Nine ADH enzymes were found in L. reuteri DSM20016, out of which 3 (PduQ, ADH6 and ADH7) belong to the group of iron-dependent enzymes that are known to transform aldehydes/ketones to alcohols. L. reuteri mutants were generated in which the three ADHs were deleted individually. The lagging growth phenotype of these deletion mutants revealed that limited NAD+/NADH recycling could be restricting their growth in the absence of ADHs. Notably, it was demonstrated that PduQ is more active in generating NAD+ during glycerol metabolism within the microcompartment by resting cells, while ADH7 functions to balance NAD+/NADH by converting 3-HPA to 1,3-PDO outside the microcompartment in the growing cells. Moreover, evaluation of ADH6 deletion mutant showed strong decrease in ethanol level, supporting the role of this bifuctional alcohol/aldehyde dehydrogenase in ethanol production. To the best of our knowledge, this is the first report revealing both internal and external recycling for cofactor homeostasis during 3-HPA conversion in L. reuteri.
Highlights
Lactobacillus species constitute an important group of lactic acid bacteria (LAB) that are normally used as probiotics, for production of fermented foods, and of biobased chemicals like lactic acid and 1,3-propanediol (1,3-PDO) [1,2,3]
Metabolic flux analysis has shown that L. reuteri uses both phosphoketolase pathway (PKP) and Embden-Meyerhof pathway (EMP) for glucose metabolism; the primary flux is through the PKP while the EMP is used as a mere shunt [10]
As 3-HPA is transported outside the microcompartment into the cytoplasm, its reduction with concomitant regeneration of NAD+ could occur either via fermentation pathways by Alcohol dehydrogenases (ADHs) and lactate dehydrogenase or instead by the
Summary
Lactobacillus species constitute an important group of lactic acid bacteria (LAB) that are normally used as probiotics, for production of fermented foods, and of biobased chemicals like lactic acid and 1,3-propanediol (1,3-PDO) [1,2,3]. The production of 1,3-PDO is achieved due to the ability of the bacteria to use glycerol as an indirect electron acceptor that helps to maintain regeneration of cofactor needed for maintaining glucose metabolism, cell growth and energy production [4,5,6,7,8]. The organism does not grow on glycerol, but addition of glycerol, 1,2-propanediol or 1,2-ethanediol to the cultivation medium induces the expression of genes in the propanediolutilization (Pdu) operon encoding shell proteins and enzymes needed for metabolism of glycerol (or the other diols) and use as electron acceptor[11]. The Pdu structural proteins form microcompartments (MCP) called metabolosomes that encapsulate the components of the metabolic pathways, and are expected to protect the cells against the toxic effect of the intermediate aldehyde, while allowing enzyme substrates (e.g. glycerol), cofactors (e.g. NAD+, NADH), and products (e.g. 1,3-PDO, 3-HP) to pass [15]
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
