AUTHOR=Otten Jonathan K. , Zou Yin , Papoutsakis Eleftherios T. 

TITLE=The potential of caproate (hexanoate) production using Clostridium kluyveri syntrophic cocultures with Clostridium acetobutylicum or Clostridium saccharolyticum

JOURNAL=Frontiers in Bioengineering and Biotechnology

VOLUME=10

YEAR=2022

URL=https://www.frontiersin.org/journals/bioengineering-and-biotechnology/articles/10.3389/fbioe.2022.965614

DOI=10.3389/fbioe.2022.965614

ISSN=2296-4185

ABSTRACT=<p>Caproate (hexanoate) and other medium-chain fatty acids are valuable platform chemicals produced by processes utilizing petroleum or plant oil. <italic>Clostridium kluyveri</italic>, growing on short chain alcohols (notably ethanol) and carboxylic acids (such as acetate) is noted for its ability to perform chain elongation to produce 4- to 8-carbon carboxylates. <italic>C. kluyveri</italic> has been studied in monoculture and coculture conditions, which lead to relatively modest carboxylate titers after long fermentation times. To assess the biosynthetic potential of <italic>C. kluyveri</italic> for caproate production from sugars through coculture fermentations, in the absence of monoculture data in the literature suitable for our coculture experiments, we first explored <italic>C. kluyveri</italic> monocultures. Some monocultures achieved caproate titers of 150 to over 200 mM in 40–50 h with a production rate of 7.9 mM/h. Based on that data, we then explored two novel, syntrophic coculture partners for producing caproate from sugars: <italic>Clostridium acetobutylicum</italic> and <italic>Clostridium saccharolyticum</italic>. Neither species has been cocultured with <italic>C. kluyveri</italic> before, and both demonstrate promising results. Our experiments of <italic>C. kluyveri</italic> monocultures and <italic>C. kluyveri</italic>—<italic>C. saccharolyticum</italic> cocultures demonstrate exceptionally high caproate titers (145–200 mM), fast production rates (3.25–8.1 mM/h), and short fermentation times (18–45 h). These results represent the most caproate produced by a <italic>C. kluyveri</italic> coculture in the shortest known fermentation time. We also explored the possibility of heterologous cell fusion between the coculture pairs similar to the results seen previously in our group with <italic>C. acetobutylicum</italic> and <italic>Clostridium ljungdahlii.</italic> Fusion events were observed only in the <italic>C. acetobutylicum</italic>—<italic>C. kluyveri</italic> coculture pair, and we offer an explanation for the lack of fusion between <italic>C. saccharolyticum</italic> and <italic>C. kluyveri</italic>. This work supports the promise of coculture biotechnology for sustainable production of caproate and other platform chemicals.</p>