Designer fungus FAD glucose dehydrogenase capable of direct electron transfer

Kohei Ito, Junko Okuda-Shimazaki, Kazushige Mori, Katsuhiro Kojima, Wakako Tsugawa, Kazunori Ikebukuro, Chi En Lin, Jeffrey LaBelle, Hiromi Yoshida, Koji Sode

Research output: Contribution to journalArticle

3 Citations (Scopus)

Abstract

Fungi-derived flavin adenine dinucleotide glucose dehydrogenases (FADGDHs) are currently the most popular and advanced enzymes for self-monitoring of blood glucose sensors; however, the achievement of direct electron transfer (DET) with FADGDHs is difficult. In this study, a designer FADGDH was constructed by fusing Aspergillus flavus derived FADGDH (AfGDH) and a Phanerochaete chrisosporium CDH (PcCDH)-derived heme b-binding cytochrome domain to develop a novel FADGDH that is capable of direct electron transfer with an electrode. A structural prediction suggested that the heme in the CDH may exist in proximity to the FAD of AfGDH if the heme b-binding cytochrome domain is fused to the AfGDH N-terminal region. Spectroscopic observations of recombinantly produced designer FADGDH confirmed the intramolecular electron transfer between FAD and the heme. A decrease in pH and the presence of a divalent cation improved the intramolecular electron transfer. An enzyme electrode with the immobilized designer FADGDH showed an increase in current immediately after the addition of glucose in a glucose concentration-dependent manner, whereas those with wild-type AfGDH did not show an increase in current. Therefore, the designer FADGDH was confirmed to be a novel GDH that possesses electrode DET ability. The difference in the surface electrostatic potentials of AfGDH and the catalytic domain of PcCDH might be why their intramolecular electron transfer ability is inferior to that of CDH. These relevant and consistent findings provide us with a novel strategic approach for the improvement of the DET properties of designer FADGDH. (241 words)

Original languageEnglish (US)
JournalBiosensors and Bioelectronics
DOIs
StateAccepted/In press - Jan 1 2018

Fingerprint

Glucose 1-Dehydrogenase
Glucose Dehydrogenases
Flavin-Adenine Dinucleotide
Fungi
Glucose
Electrons
Aspergillus flavus
Aspergillus
Heme
Cytochrome b Group
Phanerochaete
Aptitude
Cytochromes
Electrodes
Oxidoreductases
Enzyme electrodes
Blood Glucose Self-Monitoring
Glucose sensors
Proteins
Divalent Cations

Keywords

  • Cellobiose dehydrogenase
  • Designer FADGDH
  • Direct electron transfer
  • Fusion protein
  • Glucose dehydrogenase
  • Heme b-binding cytochrome domain

ASJC Scopus subject areas

  • Biotechnology
  • Biophysics
  • Biomedical Engineering
  • Electrochemistry

Cite this

Ito, K., Okuda-Shimazaki, J., Mori, K., Kojima, K., Tsugawa, W., Ikebukuro, K., ... Sode, K. (Accepted/In press). Designer fungus FAD glucose dehydrogenase capable of direct electron transfer. Biosensors and Bioelectronics. https://doi.org/10.1016/j.bios.2018.07.027

Designer fungus FAD glucose dehydrogenase capable of direct electron transfer. / Ito, Kohei; Okuda-Shimazaki, Junko; Mori, Kazushige; Kojima, Katsuhiro; Tsugawa, Wakako; Ikebukuro, Kazunori; Lin, Chi En; LaBelle, Jeffrey; Yoshida, Hiromi; Sode, Koji.

In: Biosensors and Bioelectronics, 01.01.2018.

Research output: Contribution to journalArticle

Ito, K, Okuda-Shimazaki, J, Mori, K, Kojima, K, Tsugawa, W, Ikebukuro, K, Lin, CE, LaBelle, J, Yoshida, H & Sode, K 2018, 'Designer fungus FAD glucose dehydrogenase capable of direct electron transfer', Biosensors and Bioelectronics. https://doi.org/10.1016/j.bios.2018.07.027
Ito, Kohei ; Okuda-Shimazaki, Junko ; Mori, Kazushige ; Kojima, Katsuhiro ; Tsugawa, Wakako ; Ikebukuro, Kazunori ; Lin, Chi En ; LaBelle, Jeffrey ; Yoshida, Hiromi ; Sode, Koji. / Designer fungus FAD glucose dehydrogenase capable of direct electron transfer. In: Biosensors and Bioelectronics. 2018.
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abstract = "Fungi-derived flavin adenine dinucleotide glucose dehydrogenases (FADGDHs) are currently the most popular and advanced enzymes for self-monitoring of blood glucose sensors; however, the achievement of direct electron transfer (DET) with FADGDHs is difficult. In this study, a designer FADGDH was constructed by fusing Aspergillus flavus derived FADGDH (AfGDH) and a Phanerochaete chrisosporium CDH (PcCDH)-derived heme b-binding cytochrome domain to develop a novel FADGDH that is capable of direct electron transfer with an electrode. A structural prediction suggested that the heme in the CDH may exist in proximity to the FAD of AfGDH if the heme b-binding cytochrome domain is fused to the AfGDH N-terminal region. Spectroscopic observations of recombinantly produced designer FADGDH confirmed the intramolecular electron transfer between FAD and the heme. A decrease in pH and the presence of a divalent cation improved the intramolecular electron transfer. An enzyme electrode with the immobilized designer FADGDH showed an increase in current immediately after the addition of glucose in a glucose concentration-dependent manner, whereas those with wild-type AfGDH did not show an increase in current. Therefore, the designer FADGDH was confirmed to be a novel GDH that possesses electrode DET ability. The difference in the surface electrostatic potentials of AfGDH and the catalytic domain of PcCDH might be why their intramolecular electron transfer ability is inferior to that of CDH. These relevant and consistent findings provide us with a novel strategic approach for the improvement of the DET properties of designer FADGDH. (241 words)",
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AU - Ito, Kohei

AU - Okuda-Shimazaki, Junko

AU - Mori, Kazushige

AU - Kojima, Katsuhiro

AU - Tsugawa, Wakako

AU - Ikebukuro, Kazunori

AU - Lin, Chi En

AU - LaBelle, Jeffrey

AU - Yoshida, Hiromi

AU - Sode, Koji

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N2 - Fungi-derived flavin adenine dinucleotide glucose dehydrogenases (FADGDHs) are currently the most popular and advanced enzymes for self-monitoring of blood glucose sensors; however, the achievement of direct electron transfer (DET) with FADGDHs is difficult. In this study, a designer FADGDH was constructed by fusing Aspergillus flavus derived FADGDH (AfGDH) and a Phanerochaete chrisosporium CDH (PcCDH)-derived heme b-binding cytochrome domain to develop a novel FADGDH that is capable of direct electron transfer with an electrode. A structural prediction suggested that the heme in the CDH may exist in proximity to the FAD of AfGDH if the heme b-binding cytochrome domain is fused to the AfGDH N-terminal region. Spectroscopic observations of recombinantly produced designer FADGDH confirmed the intramolecular electron transfer between FAD and the heme. A decrease in pH and the presence of a divalent cation improved the intramolecular electron transfer. An enzyme electrode with the immobilized designer FADGDH showed an increase in current immediately after the addition of glucose in a glucose concentration-dependent manner, whereas those with wild-type AfGDH did not show an increase in current. Therefore, the designer FADGDH was confirmed to be a novel GDH that possesses electrode DET ability. The difference in the surface electrostatic potentials of AfGDH and the catalytic domain of PcCDH might be why their intramolecular electron transfer ability is inferior to that of CDH. These relevant and consistent findings provide us with a novel strategic approach for the improvement of the DET properties of designer FADGDH. (241 words)

AB - Fungi-derived flavin adenine dinucleotide glucose dehydrogenases (FADGDHs) are currently the most popular and advanced enzymes for self-monitoring of blood glucose sensors; however, the achievement of direct electron transfer (DET) with FADGDHs is difficult. In this study, a designer FADGDH was constructed by fusing Aspergillus flavus derived FADGDH (AfGDH) and a Phanerochaete chrisosporium CDH (PcCDH)-derived heme b-binding cytochrome domain to develop a novel FADGDH that is capable of direct electron transfer with an electrode. A structural prediction suggested that the heme in the CDH may exist in proximity to the FAD of AfGDH if the heme b-binding cytochrome domain is fused to the AfGDH N-terminal region. Spectroscopic observations of recombinantly produced designer FADGDH confirmed the intramolecular electron transfer between FAD and the heme. A decrease in pH and the presence of a divalent cation improved the intramolecular electron transfer. An enzyme electrode with the immobilized designer FADGDH showed an increase in current immediately after the addition of glucose in a glucose concentration-dependent manner, whereas those with wild-type AfGDH did not show an increase in current. Therefore, the designer FADGDH was confirmed to be a novel GDH that possesses electrode DET ability. The difference in the surface electrostatic potentials of AfGDH and the catalytic domain of PcCDH might be why their intramolecular electron transfer ability is inferior to that of CDH. These relevant and consistent findings provide us with a novel strategic approach for the improvement of the DET properties of designer FADGDH. (241 words)

KW - Cellobiose dehydrogenase

KW - Designer FADGDH

KW - Direct electron transfer

KW - Fusion protein

KW - Glucose dehydrogenase

KW - Heme b-binding cytochrome domain

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