Cloning, purification, and characterization of a thermostable α-l-arabinofuranosidase from Anoxybacillus kestanbolensis AC26Sari

Çanakçı, Sabriye; Kacagan, Murat; İnan, Kadriye; Beldüz, Ali Osman; Saha, Badal C.

doi:10.1007/s00253-008-1584-1

Cited by 31 publications

(30 citation statements)

References 26 publications

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“…In addition, the enzymes function with maximum activity at high temperatures, up to 75°C for CpAbf51A. Comparable data (with pH 5.0 to 7.0 and temperatures of 60°C to 75°C) were also reported for arabinofuranosidases and arabinanases from other thermophilic bacteria (10,(39)(40)(41)(42)(43)(44)(45)(46)(47)(48)(49)(50). However, higher optimum temperatures from 80°C to 90°C in the same pH range were reported for enzymes from Caldicellulosiruptor saccharolyticus (46,51), Sulfolobus solfataricus (52), Clostridium thermocellum (53), and Thermotoga maritima (54).…”

Section: Discussionmentioning

confidence: 54%

Enzymatic Mechanism for Arabinan Degradation and Transport in the Thermophilic Bacterium Caldanaerobius polysaccharolyticus

Wefers

Dong

Abdelhamid

et al. 2017

Appl Environ Microbiol

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The plant cell wall polysaccharide arabinan provides an important supply of arabinose, and unraveling arabinan-degrading strategies by microbes is important for understanding its use as a source of energy. Here, we explored the arabinan-degrading enzymes in the thermophilic bacterium Caldanaerobius polysaccharolyticus and identified a gene cluster encoding two glycoside hydrolase (GH) family 51 ␣-L-arabinofuranosidases (CpAbf51A, CpAbf51B), a GH43 endoarabinanase (CpAbn43A), a GH27 ␤-L-arabinopyranosidase (CpAbp27A), and two GH127 ␤-Larabinofuranosidases (CpAbf127A, CpAbf127B). The genes were expressed as recombinant proteins, and the functions of the purified proteins were determined with para-nitrophenyl (pNP)-linked sugars and naturally occurring pectin structural elements as the substrates. The results demonstrated that CpAbn43A is an endoarabinanase while CpAbf51A and CpAbf51B are ␣-L-arabinofuranosidases that exhibit diverse substrate specificities, cleaving ␣-1,2, ␣-1,3, and ␣-1,5 linkages of purified arabinan-oligosaccharides. Furthermore, both CpAbf127A and CpAbf127B cleaved ␤-arabinofuranose residues in complex arabinan side chains, thus providing evidence of the function of this family of enzymes on such polysaccharides. The optimal temperatures of the enzymes ranged between 60°C and 75°C, and CpAbf43A and CpAbf51A worked synergistically to release arabinose from branched and debranched arabinan. Furthermore, the hydrolytic activity on branched arabinan oligosaccharides and degradation of pectic substrates by the endoarabinanase and L-arabinofuranosidases suggested a microbe equipped with diverse activities to degrade complex arabinan in the environment. Based on our functional analyses of the genes in the arabinan degradation cluster and the substrate-binding studies on a component of the cognate transporter system, we propose a model for arabinan degradation and transport by C. polysaccharolyticus. IMPORTANCE Genomic DNA sequencing and bioinformatic analysis allowed the identification of a gene cluster encoding several proteins predicted to function in arabinan degradation and transport in C. polysaccharolyticus. The analysis of the recombinant proteins yielded detailed insights into the putative arabinan metabolism of this thermophilic bacterium. The use of various branched arabinan oligosaccharides provided a detailed understanding of the substrate specificities of the enzymes and allowed assignment of two new GH127 polypeptides as ␤-L-arabinofuranosidases able to degrade pectic substrates, thus expanding our knowledge of this rare group of glycoside hydrolases. In addition, the enzymes showed synergistic effects for the degradation of arabinans at elevated temperatures. The enzymes characterized from the gene cluster are, therefore, of utility for arabinose production in both the biofuel and food industries.

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Section: Discussionmentioning

confidence: 54%

Enzymatic Mechanism for Arabinan Degradation and Transport in the Thermophilic Bacterium Caldanaerobius polysaccharolyticus

Wefers

Dong

Abdelhamid

et al. 2017

Appl Environ Microbiol

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“…Only two α‐ l ‐arabinofuranosidases, those isolated from Bifidobacterium breve (Shin et al. 2003) and A. kestanbolensis (Canakci et al. 2008), both of which exhibit narrow substrate specificity, displayed activity only for p NP‐Araf.…”

Section: Discussionmentioning

confidence: 99%

“…2006). According to the carbohydrate‐active enzyme server CAZY (http://www.cazy.org/Glycoside-Hydrolases.html), these enzymes belong to GH families 3, 10, 43, 51, 54 and 64. α‐ l ‐Arabinofuranosidases have been reported in various micro‐organisms, including the bacteria Anoxybacillus kestanbolensis (Canakci et al. 2008), Bacillus subtilis (Inacio et al.…”

Section: Introductionmentioning

confidence: 99%

Hydrolytic properties of a thermostable α-l-arabinofuranosidase from Caldicellulosiruptor saccharolyticus

Lim

Yoon

Seo

et al. 2010

Journal of Applied Microbiology

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Aims: To characterize of a thermostable recombinant α‐l‐arabinofuranosidase from Caldicellulosiruptor saccharolyticus for the hydrolysis of arabino‐oligosaccharides to l‐arabinose. Methods and Results: A recombinant α‐l‐arabinofuranosidase from C. saccharolyticus was purified by heat treatment and Hi‐Trap anion exchange chromatography with a specific activity of 28·2 U mg−1. The native enzyme was a 58‐kDa octamer with a molecular mass of 460 kDa, as measured by gel filtration. The catalytic residues and consensus sequences of the glycoside hydrolase 51 family of α‐l‐arabinofuranosidases were completely conserved in α‐l‐arabinofuranosidase from C. saccharolyticus. The maximum enzyme activity was observed at pH 5·5 and 80°C with a half‐life of 49 h at 75°C. Among aryl‐glycoside substrates, the enzyme displayed activity only for p‐nitrophenyl‐α‐l‐arabinofuranoside [maximum kcat/Km of 220 m(mol l−1)−1 s−1] and p‐nitrophenyl‐α‐l‐arabinopyranoside. This substrate specificity differs from those of other α‐l‐arabinofuranosidases. In a 1 mmol l−1 solution of each sugar, arabino‐oligosaccharides with 2–5 monomer units were completely hydrolysed to l‐arabinose within 13 h in the presence of 30 U ml−1 of enzyme at 75°C. Conclusions: The novel substrate specificity and hydrolytic properties for arabino‐oligosaccharides of α‐l‐arabinofuranosidase from C. saccharolyticus demonstrate the potential in the commercial production of l‐arabinose in concert with endoarabinanase and/or xylanase. Significance and Impact of the Study: The findings of this work contribute to the knowledge of hydrolytic properties for arabino‐oligosaccharides performed by thermostable α‐l‐arabinofuranosidase.

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“…) and Anoxybacillus kestanbolensis (Canakci et al . ), which displayed activity only for p NP‐Arf. Altogether, these results indicate that Tt‐Afs has high selectivity for the artificial substrate, pNP‐Arf.…”

Section: Discussionmentioning

confidence: 99%

Characterization of a novel arabinose-tolerantα-l-arabinofuranosidase with high ginsenoside Rc to ginsenoside Rd bioconversion productivity

Xie

Zhao

et al. 2016

J Appl Microbiol

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Cloning, purification, and characterization of a thermostable α-l-arabinofuranosidase from Anoxybacillus kestanbolensis AC26Sari

Cited by 31 publications

References 26 publications

Enzymatic Mechanism for Arabinan Degradation and Transport in the Thermophilic Bacterium Caldanaerobius polysaccharolyticus

Enzymatic Mechanism for Arabinan Degradation and Transport in the Thermophilic Bacterium Caldanaerobius polysaccharolyticus

Hydrolytic properties of a thermostable α-l-arabinofuranosidase from Caldicellulosiruptor saccharolyticus

Characterization of a novel arabinose-tolerantα-l-arabinofuranosidase with high ginsenoside Rc to ginsenoside Rd bioconversion productivity

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