Electrostatic Immobilization of D-Xylose Isomerase to a Cation Exchanger for the Conversion of D-Xylose to D-Xylulose
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Since D-xylose is not fermentable in Saccharomyces cerevisiae, its conversion to D-xylulose is required for its application in biotechnological industries using S. cerevisiae. In order to convert D-xylose to D-xylulose by way of an enzyme immobilized system, D-xylose isomerase (XI) of Escherichia coli was fused with 10-arginine tag (R10) at its C-terminus for the simple purification and immobilization process using a cation exchanger. The fusion protein XIR10 was overexpressed in recombinant E. coli and purified to a high purity by a single step of cation exchange chromatography. The purified XIR10 was immobilized to a cation exchanger via the electrostatic interaction with the C-terminal 10-arginine tag. Both the free and immobilized XIR10 exhibited similar XI activities at various pH values and temperatures, indicating that the immobilization to the cation exchanger has a small effect on the enzymatic function of XIR10. Under optimized conditions for the immobilized XIR10, Dxylose was isomerized to D-xylulose with a conversion yield of 25%. Therefore, the results of this study clearly demonstrate that the electrostatic immobilization of XIR10 via the interaction between the 10-arginine tag and a cation exchanger is an applicable form of the conversion of D-xylose to D-xylulose.
Abstract
Since D-xylose is not fermentable in Saccharomyces cerevisiae, its conversion to D-xylulose is required for its application in biotechnological industries using S. cerevisiae. In order to convert D-xylose to D-xylulose by way of an enzyme immobilized system, D-xylose isomerase (XI) of Escherichia coli was fused with 10-arginine tag (R10) at its C-terminus for the simple purification and immobilization process using a cation exchanger. The fusion protein XIR10 was overexpressed in recombinant E. coli and purified to a high purity by a single step of cation exchange chromatography. The purified XIR10 was immobilized to a cation exchanger via the electrostatic interaction with the C-terminal 10-arginine tag. Both the free and immobilized XIR10 exhibited similar XI activities at various pH values and temperatures, indicating that the immobilization to the cation exchanger has a small effect on the enzymatic function of XIR10. Under optimized conditions for the immobilized XIR10, Dxylose was isomerized to D-xylulose with a conversion yield of 25%. Therefore, the results of this study clearly demonstrate that the electrostatic immobilization of XIR10 via the interaction between the 10-arginine tag and a cation exchanger is an applicable form of the conversion of D-xylose to D-xylulose.
참고문헌 (18)
- Brewer, S. J, "The purification of recombinant proteins using C-terminal polyarginine fusions", Trends Biotechnol, 3, pp.119~122 (1985)
- Chandrakant, P, "Application of a compatible xylose isomerase in simultaneous bioconversion of glucose and xylose to ethanol", Biotechnol. Bioprocess Eng, 5, pp.32~39 (2000)
- Farber, G. K, "Crystallographic studies of the mechanism of xylose isomerase", Biochemistry, 28, pp.7289~7297 (1989)
- Gong, C, "Direct fermentation of D-xylose to ethanol by a xylose-fermentating yeast mutant, Candida sp. XF 217", Biotechnol. Lett, 3, pp.245~250 (1981)
- Jokela, J., "Isomerization of pentose and hexose sugars by an enzyme reactor packed with cross-linked xylose isomerase crystals", Enzyme Microb. Technol, 31, pp.67~76 (2002)
- Kim, S. G, "Application of poly-arginine fused minichaperone to renaturation of cyclodextrin glycosyltransferase expressed in recombinant Escherichia coli", ENZYME AND MICROBIAL TECHNOLOGY, 39, pp.459~465, ELSEVIER SCIENCE INC
- Kim, S. G, "Production and solid-phase refolding of human glucagon-like peptide-1 using recombinant Escherichia coli", PROTEIN EXPRESSION AND PURIFICATION, pp.78(2), 197~203[DOI]
- Korndorfer, I. P, "The structure of rhamnose isomerase from Escherichia coli and its relation with xylose isomerase illustrates a change between inter and intra-subunit complementation during evolution", J. Mol. Biol, 300, pp.917~933 (2000)
- Kweon, D. H, "Immobilization of Bacillus macerans cyclodextrin glycosyltransferase fused with poly-lysine using cation exchanger", Enzyme Microb. Technol, 36, pp.373~615 (2005)
- Kweon, D. H, "Purification and refolding of cyclodextrin glycosyltransferase expressed from recombinant Escherichia coli", Journal of Inclusion Phenomena and Macrocyclic Chemistry, 50, pp.37~41 (2004)
- 이대희, "Characterization of polycationic amino acids fusion systems for ion-exchange purification of cyclodextrin glycosyltransferase from recombinant Escherichia coli", BIOTECHNOLOGY PROGRESS, 18, pp.303~308 (2002), AMER CHEMICAL SOC
- Lee, D. H, "Folding machineries displayed on a cation-exchanger for the concerted refolding of cysteine-or proline-rich proteins", BMC Biotechnol, 9, pp.27~ (2009)
- Lehmacher, A, "Isolation and characterization of an extremely thermostable D-xylose isomerase from Thermus aquaticus HB 8", J. Gen. Microbiol, 136, pp.679~686 (1990)
- Mateo, C, "Improvement of enzyme activity, stability and selectivity via immobilization techniques", Enzyme Microb. Technol, 40, pp.1451~1463 (2007)
- Nock, S, "Reversible, sitespecific immobilization of polyarginine-tagged fusion proteins on mica surfaces", FEBS Lett, 414, pp.233~238 (1997)
- 이은규, "Solid-phase Refolding of Poly-lysine Tagged Fusion Protein of hEGF and Angiogenin", Biotechnology and Bioprocess Engineering, 7(1), pp.1~5 (2002), 한국생물공학회
- Pronk, J, "Preparation of D-xylulose from Dxylose", Enzyme Microb. Technol, 10, pp.537~542 (1988)
- van Maris, A, "Development of efficient xylose fermentation in Saccharomyces cerevisiae: xylose isomerase as a key component", Biofuels, 108, pp.179~204 (2007)