D-xylose (INS 967) is an important carbohydrate, which is not only one of the main hydrolysis products of cellulose in fuel ethanol, but also the perfect sweetener and therapeutical agent for diabetics. Therefore, a rapid, sensitive and selective detection of xylose would be of great interest in developing cellulose-based ethanol, pharmacology, human nutrition and food technology.To date,several methods including high performance liquid chromatography, ion chromatography have been developed to detect xylose. However, these techniques have experienced practical challenges including expensive instrument, time-consuming, complex processes of samples handling and lower insensitivity.

In collaboration with Prof. Marco Mascini from University of Florence, Italy, Prof. LIU Aihua, group leader of the Laboratory for Biosensing, Qingdao Institute of Bioenergy and Bioprocess Technology (QIBEBT), Chinese Academy of Sciences (CAS), efficiently developed a xylose dehydrogenase (XDH) bacteria surface displayed system on basis of ice nucleation protein. This work has been published in leading journal in analytical chemistry field (Analytical Chemistry 2012,84, 275-282). Research assistant LIANG Bo constructed the plasmid pTInaPbN-Xdh, which was expressed in E. coli BL21 (DE3). Most proteins were shown to exist on the outer membrane of cells, and XDH enzyme activity of outer membrane fraction accounted for 77 % of the cell lysate activity. Furthermore, the recombinant strains were more stable than the free XDH in the cytoplasm of original strain.

XDH on the surface of bacteria could convert D-xylose to D-xylonolactone with coenzyme NAD+, which is reduced to NADH, exhibiting a typical absorption peak at 340 nm. Using the bacteria display system, D-xylose could be detected with a wide linear range (5-900 μM). The presence of 100-fold excess of other sugars including cellobiose, D-fructose, D-sucrose, D-maltose, D-glucose, D-mannose, D-galactose, D-xylitol and D-ribose as well as 10-fold excess of L-arabinose were found to have no interference to the detection of D-xylose(0.1 mM).

The proposed method showed here can be applied to rapidly detect D-xylose for fermentation, bioprocess, bioenergy and food industry. The XDH-bacteria can also be used to develop electrochemical xylose biosensor with high sensitivity and selectivity (Biosensors and Bioelectronics 2012, DOI:10.1016/j.bios.2011.12.027.).

Figure 1. Construction of xylose dehydrogenase bacteria surface displayed system based on ice nucleation protein for D-xylose detection with UV-visible spectrophotometer.

This research was funded by the Knowledge Innovation Project in Biotechnology, Chinese Academy of Sciences.

Link of the publications:

http://pubs.acs.org/doi/abs/10.1021/ac202513u

http://www.sciencedirect.com/science/article/pii/S095656631100827X