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Cellobionic acid utilization: from Neurospora crassa to Saccharomyces cerevisiae.

Li X, Chomvong K, Yu VY, Liang JM, Lin Y, Cate JH - Biotechnol Biofuels (2015)

Bottom Line: Intracellular cellobionic acid was further cleaved to glucose 1-phosphate and gluconic acid by CAP.However, it remains unclear how N. crassa utilizes extracellular gluconic acid.The aldonic acid pathway was successfully implemented in Saccharomyces cerevisiae when N. crassa gluconokinase was co-expressed, resulting in cellobionic acid consumption in both aerobic and anaerobic conditions.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular and Cell Biology, University of California, Berkeley, CA USA.

ABSTRACT

Background: Economical production of fuels and chemicals from plant biomass requires the efficient use of sugars derived from the plant cell wall. Neurospora crassa, a model lignocellulosic degrading fungus, is capable of breaking down the complex structure of the plant cell wall. In addition to cellulases and hemicellulases, N. crassa secretes lytic polysaccharide monooxygenases (LPMOs), which cleave cellulose by generating oxidized sugars-particularly aldonic acids. However, the strategies N. crassa employs to utilize these sugars are unknown.

Results: We identified an aldonic acid utilization pathway in N. crassa, comprised of an extracellular hydrolase (NCU08755), cellobionic acid transporter (CBT-1, NCU05853) and cellobionic acid phosphorylase (CAP, NCU09425). Extracellular cellobionic acid could be imported directly by CBT-1 or cleaved to gluconic acid and glucose by a β-glucosidase (NCU08755) outside the cells. Intracellular cellobionic acid was further cleaved to glucose 1-phosphate and gluconic acid by CAP. However, it remains unclear how N. crassa utilizes extracellular gluconic acid. The aldonic acid pathway was successfully implemented in Saccharomyces cerevisiae when N. crassa gluconokinase was co-expressed, resulting in cellobionic acid consumption in both aerobic and anaerobic conditions.

Conclusions: We successfully identified a branched aldonic acid utilization pathway in N. crassa and transferred its essential components into S. cerevisiae, a robust industrial microorganism.

No MeSH data available.


Related in: MedlinePlus

N. crassa growth on aldonic acids. a Biomass accumulation of N. crassa provided with different carbon sources after 48 h. All samples were started with an equal inoculum of 1 × 106 cells/mL. The plate was imaged on a black background to highlight fungal growth. b Relative abundance of aldonic acids in the supernatants of cells provided with cellobionic acid at the time of inoculation (top) and 40 h (bottom). G1 glucose, G2 cellobiose, A1 gluconic acid, A2 cellobionic acid, A3 cellotrionic acid, NC no carbon control.
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Fig1: N. crassa growth on aldonic acids. a Biomass accumulation of N. crassa provided with different carbon sources after 48 h. All samples were started with an equal inoculum of 1 × 106 cells/mL. The plate was imaged on a black background to highlight fungal growth. b Relative abundance of aldonic acids in the supernatants of cells provided with cellobionic acid at the time of inoculation (top) and 40 h (bottom). G1 glucose, G2 cellobiose, A1 gluconic acid, A2 cellobionic acid, A3 cellotrionic acid, NC no carbon control.

Mentions: To test for the presence of an aldonic acid utilization pathway, N. crassa was grown aerobically on two of the simplest aldonic acids—gluconic acid and cellobionic acid. Two days after inoculation, growth on cellobionic acid was robust while that on gluconic acid was minimal (Fig. 1a). To assess N. crassa’s ability to utilize aldonic acids, supernatants of the cells grown in cellobionic acid were analyzed. Comparing the starting sample to that at 40 h, the relative abundance of cellobionic acid in the media decreased (Fig. 1b). The decrease in cellobionic acid was accompanied by the appearance of gluconic acid and a small amount of cellotrionic acid, neither of which was present at the time of inoculation (Fig. 1b). These results suggested that N. crassa was capable of processing extracellular cellobionic acid and consuming it.Fig. 1


Cellobionic acid utilization: from Neurospora crassa to Saccharomyces cerevisiae.

Li X, Chomvong K, Yu VY, Liang JM, Lin Y, Cate JH - Biotechnol Biofuels (2015)

N. crassa growth on aldonic acids. a Biomass accumulation of N. crassa provided with different carbon sources after 48 h. All samples were started with an equal inoculum of 1 × 106 cells/mL. The plate was imaged on a black background to highlight fungal growth. b Relative abundance of aldonic acids in the supernatants of cells provided with cellobionic acid at the time of inoculation (top) and 40 h (bottom). G1 glucose, G2 cellobiose, A1 gluconic acid, A2 cellobionic acid, A3 cellotrionic acid, NC no carbon control.
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4537572&req=5

Fig1: N. crassa growth on aldonic acids. a Biomass accumulation of N. crassa provided with different carbon sources after 48 h. All samples were started with an equal inoculum of 1 × 106 cells/mL. The plate was imaged on a black background to highlight fungal growth. b Relative abundance of aldonic acids in the supernatants of cells provided with cellobionic acid at the time of inoculation (top) and 40 h (bottom). G1 glucose, G2 cellobiose, A1 gluconic acid, A2 cellobionic acid, A3 cellotrionic acid, NC no carbon control.
Mentions: To test for the presence of an aldonic acid utilization pathway, N. crassa was grown aerobically on two of the simplest aldonic acids—gluconic acid and cellobionic acid. Two days after inoculation, growth on cellobionic acid was robust while that on gluconic acid was minimal (Fig. 1a). To assess N. crassa’s ability to utilize aldonic acids, supernatants of the cells grown in cellobionic acid were analyzed. Comparing the starting sample to that at 40 h, the relative abundance of cellobionic acid in the media decreased (Fig. 1b). The decrease in cellobionic acid was accompanied by the appearance of gluconic acid and a small amount of cellotrionic acid, neither of which was present at the time of inoculation (Fig. 1b). These results suggested that N. crassa was capable of processing extracellular cellobionic acid and consuming it.Fig. 1

Bottom Line: Intracellular cellobionic acid was further cleaved to glucose 1-phosphate and gluconic acid by CAP.However, it remains unclear how N. crassa utilizes extracellular gluconic acid.The aldonic acid pathway was successfully implemented in Saccharomyces cerevisiae when N. crassa gluconokinase was co-expressed, resulting in cellobionic acid consumption in both aerobic and anaerobic conditions.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular and Cell Biology, University of California, Berkeley, CA USA.

ABSTRACT

Background: Economical production of fuels and chemicals from plant biomass requires the efficient use of sugars derived from the plant cell wall. Neurospora crassa, a model lignocellulosic degrading fungus, is capable of breaking down the complex structure of the plant cell wall. In addition to cellulases and hemicellulases, N. crassa secretes lytic polysaccharide monooxygenases (LPMOs), which cleave cellulose by generating oxidized sugars-particularly aldonic acids. However, the strategies N. crassa employs to utilize these sugars are unknown.

Results: We identified an aldonic acid utilization pathway in N. crassa, comprised of an extracellular hydrolase (NCU08755), cellobionic acid transporter (CBT-1, NCU05853) and cellobionic acid phosphorylase (CAP, NCU09425). Extracellular cellobionic acid could be imported directly by CBT-1 or cleaved to gluconic acid and glucose by a β-glucosidase (NCU08755) outside the cells. Intracellular cellobionic acid was further cleaved to glucose 1-phosphate and gluconic acid by CAP. However, it remains unclear how N. crassa utilizes extracellular gluconic acid. The aldonic acid pathway was successfully implemented in Saccharomyces cerevisiae when N. crassa gluconokinase was co-expressed, resulting in cellobionic acid consumption in both aerobic and anaerobic conditions.

Conclusions: We successfully identified a branched aldonic acid utilization pathway in N. crassa and transferred its essential components into S. cerevisiae, a robust industrial microorganism.

No MeSH data available.


Related in: MedlinePlus