Lactate and acrylate metabolism by Megasphaera elsdenii under batch and steady-state conditions

Abstract

The growth of Megasphaera elsdenii on lactate with acrylate and acrylate analogues was studied under batch and steady-state conditions. Under batch conditions, lactate was converted to acetate and propionate, and acrylate was converted into propionate. Acrylate analogues 2-methyl propenoate and 3-butenoate containing a terminal double bond were similarly converted into their respective saturated acids (isobutyrate and butyrate), while crotonate and lactate analogues 3-hydroxybutyrate and (R)-2-hydroxybutyrate were not metabolized. Under carbon-limited steady-state conditions, lactate was converted to acetate and butyrate with no propionate formed. As the acrylate concentration in the feed was increased, butyrate and hydrogen formation decreased and propionate was increasingly generated, while the calculated ATP yield was unchanged. M. elsdenii metabolism differs substantially under batch and steady-state conditions. The results support the conclusion that propionate is not formed during lactate-limited steady-state growth because of the absence of this substrate to drive the formation of lactyl coenzyme A (CoA) via propionyl-CoA transferase. Acrylate and acrylate analogues are reduced under both batch and steady-state growth conditions after first being converted to thioesters via propionyl-CoA transferase. Our findings demonstrate the central role that CoA transferase activity plays in the utilization of acids by M. elsdenii and allows us to propose a modified acrylate pathway for M. elsdenii.

Fig 1

Maximum specific growth rate (●) of M. elsdenii ATCC 17753 in lactate medium containing different initial concentrations of acrylate.

Fig 2

Controlled, batch growth of M. elsdenii on lactate (■) alone generating acetate (▼), propionate (△), and butyrate (○). Cell growth is indicated by OD (●).

Fig 3

Controlled, batch growth of M. elsdenii on lactate (■) and acrylate (◇) generating acetate (▼), propionate (△), and butyrate (○). Cell growth is indicated by OD (●).

Fig 4

Molar fraction of the total substrate flux (lactate plus acrylate) to each product under carbon-limited steady-state conditions (chemostat) versus the fraction of acrylate consumed (ξ_ACR) in an acrylate and lactate mixture. Fraction butyrate (ξ_BUT, ○) = 2 × number of moles of butyrate generated/(number of moles of lactate consumed + number of moles of acrylate consumed); fraction acetate (ξ_ACE, ▼) = number of moles of acetate generated/(number of moles of lactate consumed + number of moles of acrylate consumed); fraction propionate (ξ_PRO, △) = number of moles of propionate generated/(number of moles of lactate consumed + number of moles of acrylate consumed); fraction hydrogen (ξ_H2, ●) = number of moles of propionate generated/(number of moles of lactate consumed + number of moles of acrylate consumed). The specific activity (IU/mg dry cell weight) of propionyl-CoA transferase () is also shown.

Fig 5

Proposed pathway for metabolism of acrylate and lactate into propionate, acetate, and butyrate by M. elsdenii. This pathway is extended and modified from previous models (10, 12, 29) to show acrylate consumption via propionyl-CoA transferase and the absence of butyrate kinase.