Metabolome analysis reveals a role for glyceraldehyde 3-phosphate dehydrogenase in the inhibition of C. thermocellum by ethanol

Tian, Liang; Perot, Skyler J.; Stevenson, David; Jacobson, Tyler; Lanahan, Anthony A.; Amador-Noguez, Daniel; Olson, Daniel G.; Lynd, Lee R.

doi:10.1186/s13068-017-0961-3

Title: Metabolome analysis reveals a role for glyceraldehyde 3-phosphate dehydrogenase in the inhibition of C. thermocellum by ethanol

Journal Article · Thu Nov 30 00:00:00 EST 2017 · Biotechnology for Biofuels

DOI:https://doi.org/10.1186/s13068-017-0961-3· OSTI ID:1618703

Tian, Liang; Perot, Skyler J.; Stevenson, David; Jacobson, Tyler; Lanahan, Anthony A.; Amador-Noguez, Daniel; Olson, Daniel G.; Lynd, Lee R.

Background: Clostridium thermocellum is a promising microorganism for conversion of cellulosic biomass to biofuel, without added enzymes; however, the low ethanol titer produced by strains developed thus far is an obstacle to industrial application. Results: Here, we analyzed changes in the relative concentration of intracellular metabolites in response to gradual addition of ethanol to growing cultures. For C. thermocellum, we observed that ethanol tolerance, in experiments with gradual ethanol addition, was twofold higher than previously observed in response to a stepwise increase in the ethanol concentration, and appears to be due to a mechanism other than mutation. As ethanol concentrations increased, we found accumulation of metabolites upstream of the glyceraldehyde 3-phosphate dehydrogenase (GAPDH) reaction and depletion of metabolites downstream of that reaction. This pattern was not observed in the more ethanoltolerant organism Thermoanaerobacterium saccharolyticum. We hypothesize that the Gapdh enzyme may have diferent properties in the two organisms. Our hypothesis is supported by enzyme assays showing greater sensitivity of the C. thermocellum enzyme to high levels of NADH, and by the increase in ethanol tolerance and production when the T. saccharolyticum gapdh was expressed in C. thermocellum. Conclusions: We have demonstrated that a metabolic bottleneck occurs at the GAPDH reaction when the growth of C. thermocellum is inhibited by high levels of ethanol. We then showed that this bottleneck could be relieved by expression of the gapdh gene from T. saccharolyticum. This enzyme is a promising target for future metabolic engineering work.

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Research Organization:: Oak Ridge National Laboratory (ORNL), Oak Ridge, TN (United States)

Sponsoring Organization:: USDOE Office of Science (SC)

Grant/Contract Number:: AC05-00OR22725

OSTI ID:: 1618703

Alternate ID(s):: OSTI ID: 1626990

Journal Information:: Biotechnology for Biofuels, Journal Name: Biotechnology for Biofuels Vol. 10 Journal Issue: 1; ISSN 1754-6834

Publisher:: Springer Science + Business MediaCopyright Statement

Country of Publication:: Netherlands

Language:: English

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Cited by: 23 works

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MOESM1 of Metabolome analysis reveals a role for glyceraldehyde 3-phosphate dehydrogenase in the inhibition of C. thermocellum by ethanol Tian, Liang; Perot, Skyler; Stevenson, David https://doi.org/10.6084/m9.figshare.5648617.v1 The current record is supplemented by this dataset	dataset	January 2017
MOESM2 of Metabolome analysis reveals a role for glyceraldehyde 3-phosphate dehydrogenase in the inhibition of C. thermocellum by ethanol Tian, Liang; Perot, Skyler; Stevenson, David https://doi.org/10.6084/m9.figshare.5648635.v1 The current record is supplemented by this dataset	dataset	January 2017
MOESM3 of Metabolome analysis reveals a role for glyceraldehyde 3-phosphate dehydrogenase in the inhibition of C. thermocellum by ethanol Tian, Liang; Perot, Skyler; Stevenson, David https://doi.org/10.6084/m9.figshare.5648656.v1 The current record is supplemented by this dataset	dataset	January 2017
MOESM4 of Metabolome analysis reveals a role for glyceraldehyde 3-phosphate dehydrogenase in the inhibition of C. thermocellum by ethanol Tian, Liang; Perot, Skyler; Stevenson, David https://doi.org/10.6084/m9.figshare.5648668.v1 The current record is supplemented by this image	image	January 2017
MOESM5 of Metabolome analysis reveals a role for glyceraldehyde 3-phosphate dehydrogenase in the inhibition of C. thermocellum by ethanol Tian, Liang; Perot, Skyler; Stevenson, David https://doi.org/10.6084/m9.figshare.5648686.v1 The current record is supplemented by this image	image	January 2017
Metabolome analysis reveals a role for glyceraldehyde 3-phosphate dehydrogenase in the inhibition of C. thermocellum by ethanol Tian, Liang; Perot, Skyler; Stevenson, David https://doi.org/10.6084/m9.figshare.c.3942244 The current record is supplemented by this collection	collection	January 2017
Metabolome analysis reveals a role for glyceraldehyde 3-phosphate dehydrogenase in the inhibition of C. thermocellum by ethanol Tian, Liang; Perot, Skyler; Stevenson, David https://doi.org/10.6084/m9.figshare.c.3942244.v1 The current record is supplemented by this collection	collection	January 2017
MOESM1 of Metabolome analysis reveals a role for glyceraldehyde 3-phosphate dehydrogenase in the inhibition of C. thermocellum by ethanol Tian, Liang; Perot, Skyler; Stevenson, David https://doi.org/10.6084/M9.FIGSHARE.5648617 The current record is supplemented by this dataset	dataset	January 2017
MOESM2 of Metabolome analysis reveals a role for glyceraldehyde 3-phosphate dehydrogenase in the inhibition of C. thermocellum by ethanol Tian, Liang; Perot, Skyler; Stevenson, David https://doi.org/10.6084/M9.FIGSHARE.5648635 The current record is supplemented by this dataset	dataset	January 2017
MOESM3 of Metabolome analysis reveals a role for glyceraldehyde 3-phosphate dehydrogenase in the inhibition of C. thermocellum by ethanol Tian, Liang; Perot, Skyler; Stevenson, David https://doi.org/10.6084/m9.figshare.5648656 The current record is supplemented by this dataset	dataset	January 2017
MOESM4 of Metabolome analysis reveals a role for glyceraldehyde 3-phosphate dehydrogenase in the inhibition of C. thermocellum by ethanol Tian, Liang; Perot, Skyler; Stevenson, David https://doi.org/10.6084/m9.figshare.5648668 The current record is supplemented by this image	image	January 2017
MOESM5 of Metabolome analysis reveals a role for glyceraldehyde 3-phosphate dehydrogenase in the inhibition of C. thermocellum by ethanol Tian, Liang; Perot, Skyler; Stevenson, David https://doi.org/10.6084/m9.figshare.5648686 The current record is supplemented by this image	image	January 2017