Abstract
BackgroundXylan is the major component of hemicelluloses, which are the second most abundant polysaccharides in nature, accounting for approximately one-third of all renewable organic carbon resources on earth. Efficient degradation of xylan is the prerequisite for biofuel production. Enzymatic degradation has been demonstrated to be more attractive due to low energy consumption and environmental friendliness, when compared with chemical degradation. Exo-xylanases, as a rate-limiting factor, play an important role in the xylose production. It is of great value to identify novel exo-xylanases for efficient bioconversion of xylan in biorefinery industry.ResultsA novel glycoside hydrolase (GH) family 8 reducing-end xylose-releasing exo-oligoxylanase (Rex)-encoding gene (PbRex8) was cloned from Paenibacillus barengoltzii and heterogeneously expressed in Escherichia coli. The deduced amino acid sequence of PbRex8 shared the highest identity of 74% with a Rex from Bacillus halodurans. The recombinant enzyme (PbRex8) was purified and biochemically characterized. The optimal pH and temperature of PbRex8 were 5.5 and 55 °C, respectively. PbRex8 showed prominent activity on xylooligosaccharides (XOSs), and trace activity on xylan. It also exhibited β-1,3-1,4-glucanase and xylobiase activities. The enzyme efficiently converted corncob xylan to xylose coupled with a GH family 10 endo-xylanase, with a xylose yield of 83%. The crystal structure of PbRex8 was resolved at 1.88 Å. Structural comparison suggests that Arg67 can hydrogen-bond to xylose moieties in the -1 subsite, and Asn122 and Arg253 are close to xylose moieties in the -3 subsite, the hypotheses of which were further verified by mutation analysis. In addition, Trp205, Trp132, Tyr372, Tyr277 and Tyr369 in the grove of PbRex8 were found to involve in glucooligosaccharides interactions. This is the first report on a GH family 8 Rex from P. barengoltzii.ConclusionsA novel reducing-end xylose-releasing exo-oligoxylanase suitable for xylose production from corncobs was identified, biochemically characterized and structurally elucidated. The properties of PbRex8 may make it an excellent candidate in biorefinery industries.
Highlights
Xylan is the major component of hemicelluloses, which are the second most abundant polysaccharides in nature, accounting for approximately one-third of all renewable organic carbon resources on earth
PbRex8 displayed relatively high identities with the reported glycoside hydrolase (GH) family 8 releasing exo-oligoxylanase (Rex), sharing the highest identity of 74% with the Rex from B. halodurans (BAB05824.1) [16], followed by the Rexs from P. barcinonensis (ALP73600, 53%) [19], B. adolescentis (AAO67498.1, 33%) [17], and B. intestinalis (EDV05843.1, 33%) [18] (Fig. 1), suggesting that PbRex8 should be a novel member of GH family 8 Rexs
PbRex8 was most active at pH 5.5 and 55 °C, respectively
Summary
Xylan is the major component of hemicelluloses, which are the second most abundant polysaccharides in nature, accounting for approximately one-third of all renewable organic carbon resources on earth. As the huge consumption of fossil fuels results in enormous emission of greenhouse gases, increasing attention has been focused on the consequent air pollution and global warming [1]. To alleviate this problem, biorefinery from renewable lignocellulose biomass tends to be a potential way [2]. The complete degradation of xylans requires synergistic reaction of several xylanolytic enzymes, in which xylanases (EC 3.2.1.8) and β-xylosidases (EC 3.2.1.37) play key roles [2]. Identification of novel xylanolytic enzymes suitable for high-efficient production of xylose from xylan is still of great importance
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