Coenzyme A-transferase-independent butyrate re-assimilation in Clostridium acetobutylicum—evidence from a mathematical model

Abstract

(2014) Coenzyme A-transferaseindependent butyrate re-assimilation in Clostridium acetobutylicum -evidence from a mathematical model. Applied Microbiology and Biotechnology, 98 (21 A note on versions:The version presented here may differ from the published version or from the version of record. If you wish to cite this item you are advised to consult the publisher's version. Please see the repository url above for details on accessing the published version and note that access may require a subscription.For more information, please contact eprints@nottingham.ac.uk Coenzyme A-transferase-independent butyrate re-assimilation in Clostridium acetobutylicum -Evidence from a mathematical model Abstract: The hetero-dimeric CoA-transferase CtfA/B is believed to be crucial for the metabolic transition from acidogenesis to solventogenesis in Clostridium acetobutylicum as part of the industrial-relevant acetone-butanol-ethanol (ABE) fermentation. Here, the enzyme is assumed to mediate re-assimilation of acetate and butyrate during a pH-induced metabolic shift and to faciliate the first step of acetone formation from acetoacetyl-CoA. However, recent investigations using phosphate-limited continuous cultures have questioned this common dogma.To adress the emerging experimental discrepancies, we investigated the mutant strain Cac-ctfA398s::CT using chemostat cultures. As a consequence of this mutation, the cells are unable to express functional ctfA and are thus lacking CoA-transferase activity. A mathematical model of the pH-induced metabolic shift, which was recently developed for the wild type, is used to analyse the observed behaviour of the mutant strain with a focus on re-assimilation activities for the two produced acids.Our theoretical analysis reveals that the ctfA mutant still re-assimilates butyrate, but not acetate. Based upon this finding, we conclude that C. acetobutylicum possesses a CoA-tranferase-independent butyrate uptake mechanism that is activated by decreasing pH levels. Furthermore, we observe that butanol formation is not inhibited under our experimental conditions, as suggested by previous batch culture experiments. In concordance with recent batch experiments, acetone formation is abolished in chemostat cultures using the ctfa mutant.

Towards improved butanol production through targeted genetic modification of Clostridium pasteurianum

“…However, in the rex deletion mutant butyrate is visibly reassimilated which in the pathway proposed by Malaviya et al (2012) is coupled to acetone formation via the acetoacetyl- CoA: butyrate:CoA transferase in a similar fashion to C. acetobutylicum (Jones and Woods, 1986). These findings suggest that C. pasteurianum may be able to reassimilate butyrate without the need of acetone production either through the reverse reaction of the butyrate kinase ( buk ) and the phosphotransbutyrylase ( ptb ), as previously suggested (Hüsemann and Papoutsakis, 1989, Millat et al, 2014), or via a third, as yet undescribed, mechanism proposed by Millat et al (2014). Reassimilation of butyrate without acetone production has been described in several C. acetobutylicum inactivational mutants, most notably mutants of ctfA (Millat et al, 2014), ptb (phosphotransbutrylase) and adc (acetoacetate decarboxylase) (Lehmann et al, 2012) and butyrate kinase ( buk ) gene (Desai et al, 1999).…”

“…These findings suggest that C. pasteurianum may be able to reassimilate butyrate without the need of acetone production either through the reverse reaction of the butyrate kinase ( buk ) and the phosphotransbutyrylase ( ptb ), as previously suggested (Hüsemann and Papoutsakis, 1989, Millat et al, 2014), or via a third, as yet undescribed, mechanism proposed by Millat et al (2014). Reassimilation of butyrate without acetone production has been described in several C. acetobutylicum inactivational mutants, most notably mutants of ctfA (Millat et al, 2014), ptb (phosphotransbutrylase) and adc (acetoacetate decarboxylase) (Lehmann et al, 2012) and butyrate kinase ( buk ) gene (Desai et al, 1999). The fact that C. pasteurianum does not produce acetone under any condition tested here, or elsewhere (Biebl, 2001, Pyne et al, 2016, Sandoval et al, 2015), makes this organism an ideal target for investigations of the butyrate and acetate uptake mechanisms not reliant on CtfAB.…”

Construction of a restriction-less, marker-less mutant useful for functional genomic and metabolic engineering of the biofuel producer Clostridium acetobutylicum

“…A strain with clostron inactivated ctfAB genes was previously constructed [ 21 ]. From the physiological analysis of this mutant and with the help of a mathematical model [ 22 ], it was demonstrated that butyrate was mainly reconsumed by the phosphotransbutyrylase-buyrate-kinase pathway and not by the acetoacetyl-CoA:acetate CoA-transferase in agreement with the data presented in our study.…”

Kinetic Study of Acetone-Butanol-Ethanol Fermentation in Continuous Culture

“…Haus et al [ 40 ] presented a kinetic model that considered some of the important enzymes in continuous culture and their pH dependence. The latter kinetic model was extended in [ 41 – 43 ] to include the role of cell population dynamics as well as other enzymes involved in the metabolic shift from acidogenesis to solventogenesis. These kinetic models, however, provide limited investigation into the effects of process and kinetic parameters on the systems-level fermentation.…”