Biochemical characterization of a novel exo-oligoxylanase from Paenibacillus barengoltzii suitable for monosaccharification from corncobs

Abstract

Background: 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. 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. Results: A 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. Conclusions: A 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.