Stabilization of fungi-derived recombinant FAD-dependent glucose dehydrogenase by introducing a disulfide bond

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• 作者：Genki Sakai ; Katsuhiro Kojima ; Kazushige Mori ; Yosuke Oonishi…
• 关键词：Aspergillus flavus ; Disulfide bond ; FAD ; Glucose dehydrogenase ; Site directed mutagenesis ; Thermal stability
• 刊名：Biotechnology Letters
• 出版年：2015
• 出版时间：May 2015
• 年：2015
• 卷：37
• 期：5
• 页码：1091-1099
• 全文大小：1,922 KB
• 参考文献：1. Bak, TG (1967) Studies on glucose dehydrogenase of Aspergillus oryzae. II. Purification and physical and chemical properties. Biochim Biophys Acta 139: pp. 277-293 CrossRef
  2. Bak, TG (1967) Studies on glucose dehydrogenase of Aspergillus oryzae. III. General enzymatic properties. Biochim Biophys Acta 146: pp. 317-327 CrossRef
  3. Bak, TG, Sato, R (1967) Studies on the glucose dehydrogenase of Aspergillus oryzae. I. Induction of its synthesis by rho-benzoquinone and hydroquinone. Biochim Biophys Acta 139: pp. 265-276 CrossRef
  4. Bak, TG, Sato, R (1967) Studies on glucose dehydrogenase of Aspergillus oryzae. IV. Histidyl residue as an active site. Biochim Biophys Acta 146: pp. 328-335 CrossRef
  5. Bendtsen, JD, Nielsen, H, Heijne, G, Brunak, S (2004) Improved prediction of signal peptides: signalP 3.0. J Mol Biol 340: pp. 783-795 CrossRef
  6. Bessette, PH, Aslund, F, Beckwith, J, Georgiou, G (1999) Efficient folding of proteins with multiple disulfide bonds in the Escherichia coli cytoplasm. Proc Natl Acad Sci USA 96: pp. 13703-13708 CrossRef
  7. Clark, LC, Lyons, C (1962) Electrode systems for continuous monitoring in cardiovascular surgery. Ann NY Acad Sci 102: pp. 29-45 CrossRef
  8. Fernandez, IS, Ruiz-Duenas, FJ, Santillana, E, Ferreira, P, Martinez, MJ, Martinez, AT, Romero, A (2009) Novel structural features in the GMC family of oxidoreductases revealed by the crystal structure of fungal aryl-alcohol oxidase. Acta Crystallogr D Biol Crystallogr 65: pp. 1196-1205 CrossRef
  9. Ferri, S, Kojima, K, Sode, K (2011) Review of glucose oxidases and glucose dehydrogenases: a bird’s eye view of glucose sensing enzymes. J Diabetes Sci Technol 5: pp. 1068-1076 CrossRef
  10. Gouda, MD, Singh, SA, Rao, AGA, Thakur, MS, Karanth, NG (2003) Thermal inactivation of glucose oxidase. J Biol Chem 278: pp. 24324-24333 CrossRef
  11. Hall, T (1999) BioEdit: a user-friendly biological sequence alignment editor and analysis program for Windows 95/98/NT. Nucleic Acid Symp Ser 41: pp. 95-98
  12. Hallberg, BM, Henriksson, G, Pettersson, G, Divne, C (2002) Crystal structure of the flavoprotein domain of the extracellular flavocytochrome cellobiose dehydrogenase. J Mol Biol 315: pp. 421-434 CrossRef
  13. Macheroux, P (1999) UV-visible spectroscopy as a tool to study flavoproteins. Methods Mol Biol 131: pp. 1-7
  14. Mori, K, Nakajima, M, Kojima, K, Murakami, K, Ferri, S, Sode, K (2011) Screening of Aspergillus-derived FAD-glucose dehydrogenases from fungal genome database. Biotechnol Lett 33: pp. 2255-2263 CrossRef
  15. Müller, HM (1977) Glucons?ure bildende Enzyme bei Aspergillus niger. Zentralbl Bakteriol Parasitenkd Infektionskr Hyg 132: pp. 14-24
  16. Nielsen, H, Engelbrecht, J, Brunak, S, Heijne, G (1997) Identification of prokaryotic and eukaryotic signal peptides and prediction of their cleavage sites. Protein Eng 10: pp. 1-6 CrossRef
  17. Ogura, Y (1951) Studies on the glucose dehydrogenase of Aspergillus oryzae. J Biochem 38: pp. 75-84
  18. Ogura, Y, Nagahisa, M (1937) Untersuchungen über die Atmung und die Dehydrasesysteme von Aspergillus oryzae. Bot Mag Tokyo 51: pp. 597-612 CrossRef
  19. Pettersen, EF, Goddard, TD, Huang, CC, Couch, GS, Greenblatt, DM, Men
• 刊物类别：Biomedical and Life Sciences
• 刊物主题：Life Sciences
  Microbiology
  Biotechnology
  Applied Microbiology
  Biochemistry
  
• 出版者：Springer Netherlands
• ISSN：1573-6776

文摘

Objective To improve the stability of E. coli-produced non-glycosylated fungal FAd-glucose dehydrogenase induced a disulfide bond by site-directed mutagenesis based on structural comparisons with glucose oxidases. Results The FAD-glucose dehydrogenase (GDH) mutant Val149Cys/Gly190Cys, which was constructed based on a comparison with the three dimensional structure of glucose oxidase, showed a 110?min half-life of thermal inactivation at 45?°C, which is 13-fold greater than that of the wild-type enzyme. The considerable increase in thermal stability was further supported by Eyring plot analysis. The kinetic parameters of Val149Cys/Gly190Cys (k cat?=?760?s?, Km?=?35?mM, and catalytic efficiency (k cat/Km)?=?22?s??mM?) were almost identical to those of the wild-type enzyme (k cat?=?780?s?, Km?=?35?mM, k cat/Km?=?22?s??mM?). The substrate specificity of Val149Cys/Gly190Cys is indistinguishable from that of the wild type. Conclusion The constructed mutant, Val149Cys/Gly190Cys, had significantly increased structural stability without changing the catalytic activity and kinetic parameters of FAD-GDH, including its characteristic substrate specificity.