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

Abstract

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.

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

“…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). CpAbn43A, CpAbf51A, and CpAbf51B were thermostable at high temperatures, retaining 60% of their activity after 24 h at temperatures of up to 70°C.…”

Two Distinct α- l -Arabinofuranosidases in Caldicellulosiruptor Species Drive Degradation of Arabinose-Based Polysaccharides

“…Putative AbF51 hydrolases can be identified throughout the members of the Caldicellulosiruptor genus (Table 1). A homolog (Csac_1561; 97% protein identity with AbF51) from C. saccharolyticus was shown to have AbF characteristics and exists as a homo-octamer (32). Moreover, the transcription of AbF51-like enzymes increased significantly when several Caldicellulosiruptor strains were grown on switchgrass or other plant biomass (9,33,34), indicating an important role of AbF51 on lignocellulosic degradation.…”

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

“…5c). The optimal temperature of Tt-Afs is higher than that of the a-arabinofuranosidases from T. maritima MSB8 (Miyazaki 2005), Clostridium thermocellum (Taylor et al 2006), Caldicellulosiruptor saccharolyticus (Lim et al 2010), Bacillus pumilus ARA (Pei and Shao 2008), Bacillus stearothermophilus (Shallom et al 2002) and Thermomicrobia sp. (Birgisson et al 2004), in which the optimal temperatures range from 60 to 90°C.…”

“…), Caldicellulosiruptor saccharolyticus (Lim et al . ), Bacillus pumilus ARA (Pei and Shao ), Bacillus stearothermophilus (Shallom et al . ) and Thermomicrobia sp.…”