Rapid and selective isolation of β-xylosidase through an activitybased chemical approach

Lee Chiang Lo*, Chi Yuan Chu, Yen Ru Pan, Chin Feng Wan, Yaw-Kuen Li, Jing Jer Lin

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

5 Scopus citations

Abstract

β-Xylosidase is a key enzyme in the xylanolytic system with a great potential in many biotechnological applications, especially in the food as well as the pulp and paper industries. We have developed a chemical approach for the rapid screening and isolation of β-xylosidase. Activity probe LCL-6X targeting β-xylosidase was utilized in this study. It carries a β-xylopyranosyl recognition head, a latent trapping device consisting of a 2-fluoromethylphenoxyl group, and a biotin reporter group. The biotin reporter group serves both as a readout device and as a tool for enriching the labeled proteins. LCL-6X could selectively label a model β-xylosidase from Trichoderma koningii. All other bystander proteins used in this study, including phosphorylase b, BSA, ovalbumin, carbonic anhydrase, and trypsin inhibitor, gave negligible cross-labeling effect. With the assistance of streptavidin agarose beads and mass spectrophotometry for the recovery and identification of the biotinylated proteins, we demonstrated that LCL-6X could be successfully applied to identify a bifunctional enzyme with α-L-arabinofuranosidase/β-xylosidase activity from the total protein extract of a Pichia expressing system and a prospective β-xylosidase in the culture medium of Aspergillus fumigatus. The β-xylosidase activities from numerous microbes were also screened using the LCL-6X probe. Preliminary results showed significant differences among these microbial sources and some distinct protein bands were observed. Thus, we have successfully developed a novel chemical probe that has potential applications in xylan-related research.

Original languageEnglish
Pages (from-to)197-202
Number of pages6
JournalBiotechnology Journal
Volume1
Issue number2
DOIs
StatePublished - 1 Feb 2006

Keywords

  • β-Xylosidases
  • Activity probe
  • Mechanism-based
  • Quinone methide
  • Xylan

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