Enhanced ethanol formation by Clostridium thermocellum via pyruvate decarboxylase

Jilai Zhou,Adam M. Guss,Xiaoyu Liang,Jason T. Bouvier,Liang Tian,Lee R. Lynd,Daniel G. Olson,Skyler J. Perot,Shuen Hon

doi:10.1186/s12934-017-0783-9

Jilai Zhou, Adam M. Guss + Show 7 more

Open Access

https://doi.org/10.1186/s12934-017-0783-9

Copy DOI

Abstract

BackgroundPyruvate decarboxylase (PDC) is a well-known pathway for ethanol production, but has not been demonstrated for high titer ethanol production at temperatures above 50 °C.ResultHere we examined the thermostability of eight PDCs. The purified bacterial enzymes retained 20% of activity after incubation for 30 min at 55 °C. Expression of these PDC genes, except the one from Zymomonas mobilis, improved ethanol production by Clostridium thermocellum. Ethanol production was further improved by expression of the heterologous alcohol dehydrogenase gene adhA from Thermoanaerobacterium saccharolyticum.ConclusionThe best PDC enzyme was from Acetobactor pasteurianus. A strain of C. thermocellum expressing the pdc gene from A. pasteurianus and the adhA gene from T. saccharolyticum was able to produce 21.3 g/L ethanol from 60 g/L cellulose, which is 70% of the theoretical maximum yield.

Highlights

Pyruvate decarboxylase (PDC) is a well-known pathway for ethanol production, but has not been demonstrated for high titer ethanol production at temperatures above 50 °C
[6,7,8,9], adaptive evolution [10] and gene overexpression [11,12,13,14]; consolidated bioprocessing (CBP) for ethanol production from cellulose using C. thermocellum is still not an economical process according to the target performance metrics for cost-effective production of ethanol from lignocellulose of 90% of theoretical yield and 40 g/L titer [15]
The pyruvate ferredoxin oxidoreductase (PFOR) enzyme transfers electrons from pyruvate to ferredoxin, which must be subsequently transferred to a nicotinamide cofactor (NAD+ or NADP+) in order to be used for ethanol production

Summary

Introduction

Pyruvate decarboxylase (PDC) is a well-known pathway for ethanol production, but has not been demonstrated for high titer ethanol production at temperatures above 50 °C. Plant lignocellulosic biomass represents the most abundant renewable resource on the earth which is produced at an approximate rate of 150–170 × 109 tons annually [1]. It is one of the most attractive substrates for sustainable production of second-generation biofuels and organic chemicals. Electron transfer from ferredoxin to NAD(P) + is an exergonic reaction which is frequently coupled to another endergonic reaction for energy conservation [16]. This energy conservation, comes at the price of thermodynamic driving force [17]. The ΔrG′m value of this reaction is − 0.1 kJ/mol (Fig. 1) and would require either a

Methods

Results

Conclusion