Arabinoxylo- and Arabino-Oligosaccharides-Specific α-L-Arabinofuranosidase GH51 Isozymes from the Amylolytic Yeast Saccharomycopsis fibuligera

Abstract

IntroductionL-Arabinose is a pentose sugar that can be obtained by the hydrolysis of various plant cell wall hemicellulosic polymers such as arabinoxylans and arabinans [1]. Due to the health-beneficial functions, L-arabinose and arabino-oligosaccharides (AOS) have been considered as promising candidates for low-calorie sweeteners and prebiotic materials [2][3][4]. For the enzymatic production of L-arabinose, α-L-arabinofuranosidase (ABF; E.C. 3.2.1.55) is the key player which catalyzes the exo-type hydrolysis of L-arabinose-containing polymers [5,6]. Especially, its synergistic actions with endo-α-(1,5)-L-arabinanase (ABN; E.C. 3.2.1.99) were suggested for the cost-effective L-arabinose production from sugar beet arabinan [7,8].The ABFs possess versatile hydrolyzing activities towards the terminal non-reducing α-(1,2)-, α-(1,3)-, and/or α-(1,5)-L-arabinofuranosidic linkages in various polymeric and oligomeric substrates [5,6]. Depending on their structural features, the common microbial ABFs can be mainly categorized to the members of glycoside hydrolase (GH) families 51 and 43. To date, various ABFs have been reported from mainly bacteria, as well as a few fungi and plants [5,6,9]. Based on the genome analyses, the probable gene clusters for the utilization of L-arabinose and arabinans were found from Bacillus [10, 11], Geobacillus [12], and Corynebacterium spp. [13]. Bacillus subtilis produces two intracellular exo-ABFs GH51 and two extracellular endo-ABNs GH43, which can synergistically degrade the arabinan polymers to L-arabinose [11]. On the contrary, only a limited number of eukaryotic ABFs were genetically and enzymatically characterized from the fungal microorganisms such as Penicillium [14], Aspergillus [15], Chrysosporium [16], and Aureobasidium spp. [17]. As the eukaryotic ABFs share relatively low Two genes encoding probable α-L-arabinofuranosidase (E.C. 3.2.1.55) isozymes (ABFs) with 92.3% amino acid sequence identity, ABF51A and ABF51B, were found from chromosomes 3 and 5 of Saccharomycopsis fibuligera KJJ81, an amylolytic yeast isolated from Korean wheat-based nuruk, respectively. Each open reading frame consists of 1,551 nucleotides and encodes a protein of 517 amino acids with the molecular mass of approximately 59 kDa. These isozymes share approximately 49% amino acid sequence identity with eukaryotic ABFs from filamentous fungi. The corresponding genes were cloned, functionally expressed, and purified from Escherichia coli. SfABF51 A and SfABF51 B showed the highest activities on p-nitrophenyl arabinofuranoside at 40~45 o C and pH 7.0 in sodium phosphate buffer and at 50 o C and pH 6.0 in sodium acetate buffer, respectively. These exoacting enzymes belonging to the glycoside hydrolase (GH) family 51 could hydrolyze arabinoxylooligosaccharides (AXOS) and arabino-oligosaccharides (AOS) to produce only L-arabinose, whereas they could hardly degrade any polymeric substrates including arabinans and arabinoxylans. The detailed product analyses revealed that both SfABF51 isozymes can catalyze the ver...

Hydrolysis of Arabinoxylo-oligosaccharides by α-L-Arabinofuranosidases and β-D-Xylosidase from Bifidobacterium dentium

“…infantis possesses the considerable activities towards p -nitrophenyl-β-D-glucoside as well as p -NPXp and p -NPAf [ 27 ]. Recently, AXOS- and AOS-specific ABFs GH51 were functionally characterized from an amylolytic yeast, Saccharomycopsis fibuligera [ 28 ]. Although these ABFs similarly hydrolyzed only oligomeric substrates, not polymeric substrates, they showed no significant amino acid identities with BfdABF1 and 3.…”

Functional Characterization of Endo- and Exo-Hydrolase Genes in Arabinan Degradation Gene Cluster of Bifidobacterium longum subsp. suis

“…suis ATCC 27533 (= DSM 20211 = KCTC 3229) was obtained from Korean Collection for Type Cultures (KCTC, Daejeon, Republic of Korea). The plasmid vectors, pHCXHD [ 59 ] and pET-21a(+) (Novagen, Bagsværd, Denmark), were utilized for constitutive and IPTG-inducible gene expressions, respectively.…”

“…These enzymes belong to the Glycoside Hydrolase (GH) 2, 3, 43, 51, 54, and 62 families [ , accessed on 10 January 2024], which catalyze the hydrolysis of terminal non-reducing α-1,2-, α-1,3-, and/or α-1,5-arabinofuranoside residues from AOS and arabinans. Most ABFs have been found in various bacteria, such as Cellvibrio [ 17 ], Streptomyces [ 18 ], Thermotoga [ 19 ], Lactobacillus [ 20 ], Bacillus [ 21 , 22 ], Caldicellulosiruptor [ 23 ], Geobacillus [ 24 ], Clostridium [ 25 ], Bifidobacterium [ 26 , 27 ], and Weissella [ 28 ] species and so on, as well as from fungi and yeasts [ 29 , 30 ]. Meanwhile, endo-α-1,5- l -arabinanases (ABNs; EC 3.2.1.99) belong to the GH43 family and specifically hydrolyze arabinans into short-chain AOS.…”