Limits...
A novel bifunctional GH51 exo-α-l-arabinofuranosidase/endo-xylanase from Alicyclobacillus sp. A4 with significant biomass-degrading capacity.

Yang W, Bai Y, Yang P, Luo H, Huang H, Meng K, Shi P, Wang Y, Yao B - Biotechnol Biofuels (2015)

Bottom Line: Moreover, Ac-Abf51A showed greater synergistic effect in combination with xylanase (2.92-fold) on wheat arabinoxylan degradation than other reported enzymes, for the amounts of arabinose, xylose, and xylobiose were all increased in comparison to that by the enzymes acting individually.It was stable over a broad pH range and at high temperature, and showed greater synergistic effect with xylanase on the degradation of wheat arabinoxylan than other counterparts.The distinguished synergy might be ascribed to its bifunctional α-l-arabinofuranosidase/xylanase activity, which may represent a possible way to degrade biomass at lower enzyme loadings.

View Article: PubMed Central - PubMed

Affiliation: Key Laboratory for Feed Biotechnology of the Ministry of Agriculture, Feed Research Institute, Chinese Academy of Agricultural Sciences, No. 12 Zhongguancun South Street, Beijing, 100081 People's Republic of China.

ABSTRACT

Background: Improving the hydrolytic performance of xylanolytic enzymes on arabinoxylan is of importance in the ethanol fermentation industry. Supplementation of debranching (arabinofuranosidase) and depolymerizing (xylanase) enzymes is a way to address the problem. In the present study, we identified a bifunctional α-l-arabinofuranosidase/endo-xylanase (Ac-Abf51A) of glycoside hydrolase family 51 in Alicyclobacillus sp. strain A4. Its biochemical stability and great hydrolysis efficiency against complex biomass make it a potential candidate for the production of biofuels.

Results: The gene encoding Ac-Abf51A was cloned. The comparison of its sequence with reference proteins having resolved 3D-structures revealed nine key residues involved in catalysis and substrate-binding interaction. Recombinant Ac-Abf51A produced in Escherichia coli showed optimal activity at pH 6.0 and 60 °C with 4-nitrophenyl-α-l-arabinofuranoside as the substrate. The enzyme exhibited an exo-type mode of action on polyarabinosides by catalyzing the cleavage of α-1,2- and α-1,3-linked arabinofuranose side chains in sugar beet arabinan and water-soluble wheat arabinoxylan and α-1,5-linked arabinofuranosidic bonds in debranched sugar beet arabinan. Surprisingly, it had capacity to release xylobiose and xylotriose from wheat arabinoxylan and was active on xylooligosaccharides (xylohexaose 1.2/mM/min, xylopentaose 6.9/mM/min, and xylotetraose 19.7/mM/min), however a lower level of activity. Moreover, Ac-Abf51A showed greater synergistic effect in combination with xylanase (2.92-fold) on wheat arabinoxylan degradation than other reported enzymes, for the amounts of arabinose, xylose, and xylobiose were all increased in comparison to that by the enzymes acting individually.

Conclusions: This study for the first time reports a GH51 enzyme with both exo-α-l-arabinofuranosidase and endo-xylanase activities. It was stable over a broad pH range and at high temperature, and showed greater synergistic effect with xylanase on the degradation of wheat arabinoxylan than other counterparts. The distinguished synergy might be ascribed to its bifunctional α-l-arabinofuranosidase/xylanase activity, which may represent a possible way to degrade biomass at lower enzyme loadings.

No MeSH data available.


Related in: MedlinePlus

Time course of hydrolysis of soluble wheat arabinoxylan by Ac-Abf51A. 1, the control of substrate incubated without enzyme for 24 h; 2–5, the hydrolysate with enzyme treatment for 1 h, 6 h, 12 h, and 24 h, respectively; 6, the xylooligosaccharide standards: Ara arabinose, X1 xylose, X2 xylobiose, X3 xylotriose, X4 xylotetraose
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

License 1 - License 2
getmorefigures.php?uid=PMC4666033&req=5

Fig4: Time course of hydrolysis of soluble wheat arabinoxylan by Ac-Abf51A. 1, the control of substrate incubated without enzyme for 24 h; 2–5, the hydrolysate with enzyme treatment for 1 h, 6 h, 12 h, and 24 h, respectively; 6, the xylooligosaccharide standards: Ara arabinose, X1 xylose, X2 xylobiose, X3 xylotriose, X4 xylotetraose

Mentions: Further time course analysis of the water-soluble wheat arabinoxylan digestion by Ac-Abf51A indicated that arabinose was yielded as the sole product at the beginning, and xylobiose, xylotriose, and xylotetraose were also released after a prolonged incubation (Fig. 4). When using xylooligosaccharides as the substrates, Ac-Abf51A exhibited an endo-mode of action on xylohexaose, xylopentaose and xylotetraose, releasing products of lower degree of polymerization. However, it showed barely detectable activity on xylotriose and xylobiose. The kcat/Km values of Ac-Abf51A on xylotetraose, xylopentaose, and xylohexaose were 1.2/mM/min, 6.9/mM/min, and 19.7/mM/min, respectively (Additional file 3).Fig. 4


A novel bifunctional GH51 exo-α-l-arabinofuranosidase/endo-xylanase from Alicyclobacillus sp. A4 with significant biomass-degrading capacity.

Yang W, Bai Y, Yang P, Luo H, Huang H, Meng K, Shi P, Wang Y, Yao B - Biotechnol Biofuels (2015)

Time course of hydrolysis of soluble wheat arabinoxylan by Ac-Abf51A. 1, the control of substrate incubated without enzyme for 24 h; 2–5, the hydrolysate with enzyme treatment for 1 h, 6 h, 12 h, and 24 h, respectively; 6, the xylooligosaccharide standards: Ara arabinose, X1 xylose, X2 xylobiose, X3 xylotriose, X4 xylotetraose
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4666033&req=5

Fig4: Time course of hydrolysis of soluble wheat arabinoxylan by Ac-Abf51A. 1, the control of substrate incubated without enzyme for 24 h; 2–5, the hydrolysate with enzyme treatment for 1 h, 6 h, 12 h, and 24 h, respectively; 6, the xylooligosaccharide standards: Ara arabinose, X1 xylose, X2 xylobiose, X3 xylotriose, X4 xylotetraose
Mentions: Further time course analysis of the water-soluble wheat arabinoxylan digestion by Ac-Abf51A indicated that arabinose was yielded as the sole product at the beginning, and xylobiose, xylotriose, and xylotetraose were also released after a prolonged incubation (Fig. 4). When using xylooligosaccharides as the substrates, Ac-Abf51A exhibited an endo-mode of action on xylohexaose, xylopentaose and xylotetraose, releasing products of lower degree of polymerization. However, it showed barely detectable activity on xylotriose and xylobiose. The kcat/Km values of Ac-Abf51A on xylotetraose, xylopentaose, and xylohexaose were 1.2/mM/min, 6.9/mM/min, and 19.7/mM/min, respectively (Additional file 3).Fig. 4

Bottom Line: Moreover, Ac-Abf51A showed greater synergistic effect in combination with xylanase (2.92-fold) on wheat arabinoxylan degradation than other reported enzymes, for the amounts of arabinose, xylose, and xylobiose were all increased in comparison to that by the enzymes acting individually.It was stable over a broad pH range and at high temperature, and showed greater synergistic effect with xylanase on the degradation of wheat arabinoxylan than other counterparts.The distinguished synergy might be ascribed to its bifunctional α-l-arabinofuranosidase/xylanase activity, which may represent a possible way to degrade biomass at lower enzyme loadings.

View Article: PubMed Central - PubMed

Affiliation: Key Laboratory for Feed Biotechnology of the Ministry of Agriculture, Feed Research Institute, Chinese Academy of Agricultural Sciences, No. 12 Zhongguancun South Street, Beijing, 100081 People's Republic of China.

ABSTRACT

Background: Improving the hydrolytic performance of xylanolytic enzymes on arabinoxylan is of importance in the ethanol fermentation industry. Supplementation of debranching (arabinofuranosidase) and depolymerizing (xylanase) enzymes is a way to address the problem. In the present study, we identified a bifunctional α-l-arabinofuranosidase/endo-xylanase (Ac-Abf51A) of glycoside hydrolase family 51 in Alicyclobacillus sp. strain A4. Its biochemical stability and great hydrolysis efficiency against complex biomass make it a potential candidate for the production of biofuels.

Results: The gene encoding Ac-Abf51A was cloned. The comparison of its sequence with reference proteins having resolved 3D-structures revealed nine key residues involved in catalysis and substrate-binding interaction. Recombinant Ac-Abf51A produced in Escherichia coli showed optimal activity at pH 6.0 and 60 °C with 4-nitrophenyl-α-l-arabinofuranoside as the substrate. The enzyme exhibited an exo-type mode of action on polyarabinosides by catalyzing the cleavage of α-1,2- and α-1,3-linked arabinofuranose side chains in sugar beet arabinan and water-soluble wheat arabinoxylan and α-1,5-linked arabinofuranosidic bonds in debranched sugar beet arabinan. Surprisingly, it had capacity to release xylobiose and xylotriose from wheat arabinoxylan and was active on xylooligosaccharides (xylohexaose 1.2/mM/min, xylopentaose 6.9/mM/min, and xylotetraose 19.7/mM/min), however a lower level of activity. Moreover, Ac-Abf51A showed greater synergistic effect in combination with xylanase (2.92-fold) on wheat arabinoxylan degradation than other reported enzymes, for the amounts of arabinose, xylose, and xylobiose were all increased in comparison to that by the enzymes acting individually.

Conclusions: This study for the first time reports a GH51 enzyme with both exo-α-l-arabinofuranosidase and endo-xylanase activities. It was stable over a broad pH range and at high temperature, and showed greater synergistic effect with xylanase on the degradation of wheat arabinoxylan than other counterparts. The distinguished synergy might be ascribed to its bifunctional α-l-arabinofuranosidase/xylanase activity, which may represent a possible way to degrade biomass at lower enzyme loadings.

No MeSH data available.


Related in: MedlinePlus