Display of Glucose Oxidase on <i>Bacillus subtilis</i> Spore Surface and Its Application in Electrode

Wenhu Chen; Runlong Ma; Ping Lin; Haibo Yuan; Hongling Liu; Yi Jiang; Di Huang; Yaohong Ma; Tengfei Wang

doi:doi:10.11648/j.wjac.20220702.11

| Peer-Reviewed

Display of Glucose Oxidase on Bacillus subtilis Spore Surface and Its Application in Electrode

Wenhu Chen, Runlong Ma, Ping Lin, Haibo Yuan, Hongling Liu, Yi Jiang, Di Huang, Yaohong Ma, Tengfei Wang

Published in World Journal of Applied Chemistry (Volume 7, Issue 2)

Received: 12 March 2022 Accepted: 30 March 2022 Published: 8 April 2022

Views: Downloads:

Download PDF

Share This Article

Twitter
Linked In
Facebook

Abstract

Glucose oxidase (GOD) is widely used in the fields of food, chemistry and medicine due to its specific catalytic activity, and it is the main tool enzyme in the biological field. The biological preparation of GOD can be realized by the method of surface display on Bacillus subtilis spores. In this study, we analyzed the display characteristics of eleven cot proteins on the spore coat and determined the fluorescence intensity of different spore coat proteins by flow cytometry as follows: CotX > CotY > CotF > CotC > CotE > CotG > CotV > CotB > CotM > CotA > CotW. Next, CotC, CotX and CotY were selected to display GOD on the surface of spores, and CotX demonstrated the highest GOD activity. Subsequently, graphene oxide was added to the glassy carbon electrode and prussian blue was deposited, the surface of this electrode was fixed with spore–GOD and finally covered with a layer of Nafion solution to prepare glucose detection electrode. In the range of 0.1-7.0 mmol/L glucose, the cyclic voltammetry curve on the enzyme electrode showed a linear relationship with the glucose concentration. The calibration curve equation was I = 1.3047C_glucose+ 3.639 (R² = 0.9929), and its detection limit was 7.5 μmol/L (S/N = 3). This modified electrode has good conductivity, stability, reproducibility and can be used for the analysis and determination of glucose.

Published in	World Journal of Applied Chemistry (Volume 7, Issue 2)
DOI	10.11648/j.wjac.20220702.11
Page(s)	34-47
Creative Commons	This is an Open Access article, distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution and reproduction in any medium or format, provided the original work is properly cited.
Copyright	Copyright © The Author(s), 2024. Published by Science Publishing Group

Keywords

Bacillus subtilis, Spores, Coat Protein, Surface Display, Glucose Oxidase, Enzyme Electrode

References

[1]	Chen H, Chen Z, Wu B, Ullah J, Zhang T, Jia J, et al. (2017). Influences of Various Peptide Linkers on the Thermotoga maritima MSB8 Nitrilase Displayed on the Spore Surface of Bacillus subtilis. Microbial Physiology. 27: 64-71.
[2]	Higgins D, Dworkin J. (2012). Recent progress in Bacillus subtilis sporulation. FEMS microbiology reviews. 36: 131-148.
[3]	Wang H, Wang Y, Yang R. (2017). Recent progress in Bacillus subtilis spore-surface display: concept, progress, and future. Applied Microbiology and Biotechnology. 101: 933-949.
[4]	Setlow P. (2014). Germination of Spores of Bacillus Species: What We Know and Do Not Know. Journal of Bacteriology. 196: 1297-1305.
[5]	Liu H, Qiao H, Krajcikova D, Zhang Z, Wang H, Barak I, et al. (2016). Physical interaction and assembly of Bacillus subtilis spore coat proteins CotE and CotZ studied by atomic force microscopy. Journal of Structural Biology. 195: 245-251.
[6]	McKenney PT, Driks A, Eichenberger P. (2013). The Bacillus subtilis endospore: assembly and functions of the multilayered coat. Nature Reviews Microbiology. 11: 33-44.
[7]	Zhang G, An Y, Hossain MZ, Guo Q, Miaomiao Y, Yuan J, et al. (2019). Bacillus subtilis Spore Surface Display Technology: A Review of Its Development and Applications. J. Microbiol. Biotechnol. 29: 179-190.
[8]	Lin P, Yuan H, Du J, Liu K, Liu H, Wang T. (2020). Progress in research and application development of surface display technology using Bacillus subtilis spores. Applied Microbiology and Biotechnology. 104: 2319-2331.
[9]	Wang N, Chang C, Yao Q, Li G, Qin L, Chen L, et al. (2011). Display of Bombyx mori Alcohol Dehydrogenases on the Bacillus subtilis Spore Surface to Enhance Enzymatic Activity under Adverse Conditions. PLOS ONE. 6: e21454.
[10]	Kim J, Schumann W. (2009). Display of proteins on Bacillus subtilis endospores. Cellular and Molecular Life Sciences. 66: 3127-3136.
[11]	Svendsen A, Kiefer HV, Pedersen HB, Bochenkova AV, Andersen LH. (2017). Origin of the Intrinsic Fluorescence of the Green Fluorescent Protein. Journal of the American Chemical Society. 139: 8766-8771.
[12]	Dawidziak-Pakula A, Krasowska J, Wielgus-Kutrowska B. (2020). Analytical ultracentrifugation as a tool in the studies of aggregation of the fluorescent marker, Enhanced Green Fluorescent Protein. Acta Biochimica Polonica. 67: 85-91.
[13]	Mano N. (2019). Engineering glucose oxidase for bioelectrochemical applications. Bioelectrochemistry. 128: 218-240.
[14]	Li L, Liang B, Shi J, Li F, Mascini M, Liu A. (2012). A selective and sensitive d-xylose electrochemical biosensor based on xylose dehydrogenase displayed on the surface of bacteria and multi-walled carbon nanotubes modified electrode. Biosensors and Bioelectronics. 33: 100-105.
[15]	Hoarau M, Badieyan S, Marsh ENG. (2017). Immobilized enzymes: understanding enzyme – surface interactions at the molecular level. Organic & Biomolecular Chemistry. 15: 9539-9551.
[16]	Eda G, Chhowalla M. (2010). Chemically Derived Graphene Oxide: Towards Large-Area Thin-Film Electronics and Optoelectronics. Advanced materials. 22: 2392-2415.
[17]	Sivashankari PR, Prabaharan M. (2020). Three-dimensional porous scaffolds based on agarose/chitosan/graphene oxide composite for tissue engineering. International Journal of Biological Macromolecules. 146: 222-231.
[18]	Zhu Y, Murali S, Cai W, Li X, Suk JW, Potts JR, et al. (2010). Graphene and Graphene Oxide: Synthesis, Properties, and Applications. Advanced Materials. 22: 3906-3924.
[19]	Huang J, Lu S, Fang X, Yang Z, Liu X, Li S, et al. (2019). Optimized deposition time boosts the performance of Prussian blue modified nanoporous gold electrodes for hydrogen peroxide monitoring. Nanotechnology. 31: 045501.
[20]	Zhang M, Hou C, Halder A, Ulstrup J, Chi Q. (2017). Interlocked graphene–Prussian blue hybrid composites enable multifunctional electrochemical applications. Biosensors and Bioelectronics. 89: 570-577.
[21]	Guo Q, An Y, Yun J, Yang M, Magocha TA, Zhu J, et al. (2018). Enhanced d-tagatose production by spore surface-displayed l-arabinose isomerase from isolated Lactobacillus brevis PC16 and biotransformation. Bioresource Technology. 247: 940-946.
[22]	Jiang H, Bian Q, Zeng W, Ren P, Sun H, Lin Z, et al. (2019). Oral delivery of Bacillus subtilis spores expressing grass carp reovirus VP4 protein produces protection against grass carp reovirus infection. Fish & Shellfish Immunology. 84: 768-780.
[23]	Dai X, Liu M, Pan K, Yang J. (2018). Surface display of OmpC of Salmonella serovar Pullorum on Bacillus subtilis spores. PLOS ONE. 13: e0191627.
[24]	Tongbu L, Xinyu P, Huiying Y, Liangnian J. (1996). The production of glucose oxidase using the waste myceliums of Aspergillus niger and the effects of metal ions on the activity of glucose oxidase. Enzyme and Microbial Technology. 19: 339-342.
[25]	Wang H, Lang Q, Li L, Liang B, Tang X, Kong L, et al. (2013). Yeast Surface Displaying Glucose Oxidase as Whole-Cell Biocatalyst: Construction, Characterization, and Its Electrochemical Glucose Sensing Application. Analytical Chemistry. 85: 6107-6112.
[26]	Zheng L, Ma H, Ma Y, Meng Q, Yang J, Wang B, et al. (2019). Development and Evaluation of a Portable Electrochemical Biosensor for Detecting Uric Acid in Urine. International Journal of Electrochemical Science. 14: 9573-9583.
[27]	Koskun Y, Şavk A, Şen B, Şen F. (2018). Highly sensitive glucose sensor based on monodisperse palladium nickel/activated carbon nanocomposites. Analytica Chimica Acta. 1010: 37-43.
[28]	Vilian ATE, Chen S-M, Ali MA, Al-Hemaid FMA. (2014). Direct electrochemistry of glucose oxidase immobilized on ZrO₂ nanoparticles-decorated reduced graphene oxide sheets for a glucose biosensor. RSC Advances. 4: 30358-30367.
[29]	Haque Su, Nasar A, Inamuddin, Rahman MM. (2020). Applications of chitosan (CHI)-reduced graphene oxide (rGO)-polyaniline (PAni) conducting composite electrode for energy generation in glucose biofuel cell. Scientific Reports. 10: 10428.
[30]	Hwang B-Y, Kim B-G, Kim J-H. (2011). Bacterial Surface Display of a Co-Factor Containing Enzyme, ω-Transaminase from Vibrio fluvialis Using the Bacillus subtilis Spore Display System. Bioscience, Biotechnology, and Biochemistry. 75: 1862-1865.
[31]	Rostami A, Hinc K, Goshadrou F, Shali A, Bayat M, Hassanzadeh M, et al. (2017). Display of B. pumilus chitinase on the surface of B. subtilis spore as a potential biopesticide. Pesticide Biochemistry and Physiology. 140: 17-23.
[32]	Wang H, Yang R, Hua X, Zhang W, Zhao W. (2016). An approach for lactulose production using the CotX-mediated spore-displayed β-galactosidase as a biocatalyst. Journal of Microbiology Biotechnology. 26: 1267-1277.
[33]	Kim J-H, Roh C-H, Lee C-W, Kyung D-H, Choi S-K, Jung H-C, et al. (2007). Bacterial surface display of GFP_UV on Bacillus subtilis spores. Journal of microbiology biotechnology. 17: 677-680.
[34]	Li Q, Ning D, Wu C. (2010). Surface display of GFP using CotX as a molecular vector on Bacillus subtilis spores. Sheng wu gong cheng xue bao= Chinese journal of biotechnology. 26: 264-269.
[35]	Wang Y, Wang J, Leng F, Ma J, Bagadi A. (2020). Expression of Aspergillus niger glucose oxidase in Pichia pastoris and its antimicrobial activity against Agrobacterium and Escherichia coli. PeerJ. 8: e9010.
[36]	Khadivi Derakshan F, Darvishi F, Dezfulian M, Madzak C. (2017). Expression and Characterization of Glucose Oxidase from Aspergillus niger in Yarrowia lipolytica. Molecular Biotechnology. 59: 307-314.
[37]	Malherbe DF, du Toit M, Cordero Otero RR, van Rensburg P, Pretorius IS. (2003). Expression of the Aspergillus niger glucose oxidase gene in Saccharomyces cerevisiae and its potential applications in wine production. Applied Microbiology and Biotechnology. 61: 502-511.
[38]	Witt S, Singh M, Kalisz HM. (1998). Structural and kinetic properties of nonglycosylated recombinant Penicillium amagasakiense glucose oxidase expressed in Escherichia coli. Applied Environmental Microbiology. 64: 1405-1411.
[39]	Zhang L, Yuan S-m, Yang L-m, Fang Z, Zhao G-c. (2013). An enzymatic glucose biosensor based on a glassy carbon electrode modified with manganese dioxide nanowires. Microchimica Acta. 180: 627-633.
[40]	Wang JX, Sun XW, Wei A, Lei Y, Cai XP, Li CM, et al. (2006). Zinc oxide nanocomb biosensor for glucose detection. Applied Physics Letters. 88: 233106.

Cite This Article

Plain Text BibTeX RIS

APA Style

Wenhu Chen, Runlong Ma, Ping Lin, Haibo Yuan, Hongling Liu, et al. (2022). Display of Glucose Oxidase on Bacillus subtilis Spore Surface and Its Application in Electrode. World Journal of Applied Chemistry, 7(2), 34-47. https://doi.org/10.11648/j.wjac.20220702.11

Copy | Download

ACS Style

Wenhu Chen; Runlong Ma; Ping Lin; Haibo Yuan; Hongling Liu, et al. Display of Glucose Oxidase on Bacillus subtilis Spore Surface and Its Application in Electrode. World J. Appl. Chem. 2022, 7(2), 34-47. doi: 10.11648/j.wjac.20220702.11

Copy | Download

AMA Style

Wenhu Chen, Runlong Ma, Ping Lin, Haibo Yuan, Hongling Liu, et al. Display of Glucose Oxidase on Bacillus subtilis Spore Surface and Its Application in Electrode. World J Appl Chem. 2022;7(2):34-47. doi: 10.11648/j.wjac.20220702.11

Copy | Download

@article{10.11648/j.wjac.20220702.11,
  author = {Wenhu Chen and Runlong Ma and Ping Lin and Haibo Yuan and Hongling Liu and Yi Jiang and Di Huang and Yaohong Ma and Tengfei Wang},
  title = {Display of Glucose Oxidase on Bacillus subtilis Spore Surface and Its Application in Electrode},
  journal = {World Journal of Applied Chemistry},
  volume = {7},
  number = {2},
  pages = {34-47},
  doi = {10.11648/j.wjac.20220702.11},
  url = {https://doi.org/10.11648/j.wjac.20220702.11},
  eprint = {https://article.sciencepublishinggroup.com/pdf/10.11648.j.wjac.20220702.11},
  abstract = {Glucose oxidase (GOD) is widely used in the fields of food, chemistry and medicine due to its specific catalytic activity, and it is the main tool enzyme in the biological field. The biological preparation of GOD can be realized by the method of surface display on Bacillus subtilis spores. In this study, we analyzed the display characteristics of eleven cot proteins on the spore coat and determined the fluorescence intensity of different spore coat proteins by flow cytometry as follows: CotX > CotY > CotF > CotC > CotE > CotG > CotV > CotB > CotM > CotA > CotW. Next, CotC, CotX and CotY were selected to display GOD on the surface of spores, and CotX demonstrated the highest GOD activity. Subsequently, graphene oxide was added to the glassy carbon electrode and prussian blue was deposited, the surface of this electrode was fixed with spore–GOD and finally covered with a layer of Nafion solution to prepare glucose detection electrode. In the range of 0.1-7.0 mmol/L glucose, the cyclic voltammetry curve on the enzyme electrode showed a linear relationship with the glucose concentration. The calibration curve equation was I = 1.3047Cglucose + 3.639 (R2 = 0.9929), and its detection limit was 7.5 μmol/L (S/N = 3). This modified electrode has good conductivity, stability, reproducibility and can be used for the analysis and determination of glucose.},
 year = {2022}
}

Copy | Download

TY  - JOUR
T1  - Display of Glucose Oxidase on Bacillus subtilis Spore Surface and Its Application in Electrode
AU  - Wenhu Chen
AU  - Runlong Ma
AU  - Ping Lin
AU  - Haibo Yuan
AU  - Hongling Liu
AU  - Yi Jiang
AU  - Di Huang
AU  - Yaohong Ma
AU  - Tengfei Wang
Y1  - 2022/04/08
PY  - 2022
N1  - https://doi.org/10.11648/j.wjac.20220702.11
DO  - 10.11648/j.wjac.20220702.11
T2  - World Journal of Applied Chemistry
JF  - World Journal of Applied Chemistry
JO  - World Journal of Applied Chemistry
SP  - 34
EP  - 47
PB  - Science Publishing Group
SN  - 2637-5982
UR  - https://doi.org/10.11648/j.wjac.20220702.11
AB  - Glucose oxidase (GOD) is widely used in the fields of food, chemistry and medicine due to its specific catalytic activity, and it is the main tool enzyme in the biological field. The biological preparation of GOD can be realized by the method of surface display on Bacillus subtilis spores. In this study, we analyzed the display characteristics of eleven cot proteins on the spore coat and determined the fluorescence intensity of different spore coat proteins by flow cytometry as follows: CotX > CotY > CotF > CotC > CotE > CotG > CotV > CotB > CotM > CotA > CotW. Next, CotC, CotX and CotY were selected to display GOD on the surface of spores, and CotX demonstrated the highest GOD activity. Subsequently, graphene oxide was added to the glassy carbon electrode and prussian blue was deposited, the surface of this electrode was fixed with spore–GOD and finally covered with a layer of Nafion solution to prepare glucose detection electrode. In the range of 0.1-7.0 mmol/L glucose, the cyclic voltammetry curve on the enzyme electrode showed a linear relationship with the glucose concentration. The calibration curve equation was I = 1.3047Cglucose + 3.639 (R2 = 0.9929), and its detection limit was 7.5 μmol/L (S/N = 3). This modified electrode has good conductivity, stability, reproducibility and can be used for the analysis and determination of glucose.
VL  - 7
IS  - 2
ER  -

Copy | Download

Author Information

Wenhu Chen

State Key Laboratory of Biobased Material and Green Papermaking (LBMP), Qilu University of Technology (Shandong Academy of Sciences), Jinan, People’s Republic of China
Runlong Ma

State Key Laboratory of Biobased Material and Green Papermaking (LBMP), Qilu University of Technology (Shandong Academy of Sciences), Jinan, People’s Republic of China
Ping Lin

State Key Laboratory of Biobased Material and Green Papermaking (LBMP), Qilu University of Technology (Shandong Academy of Sciences), Jinan, People’s Republic of China
Haibo Yuan

State Key Laboratory of Biobased Material and Green Papermaking (LBMP), Qilu University of Technology (Shandong Academy of Sciences), Jinan, People’s Republic of China
Hongling Liu

State Key Laboratory of Biobased Material and Green Papermaking (LBMP), Qilu University of Technology (Shandong Academy of Sciences), Jinan, People’s Republic of China
Yi Jiang

State Key Laboratory of Biobased Material and Green Papermaking (LBMP), Qilu University of Technology (Shandong Academy of Sciences), Jinan, People’s Republic of China
Di Huang

State Key Laboratory of Biobased Material and Green Papermaking (LBMP), Qilu University of Technology (Shandong Academy of Sciences), Jinan, People’s Republic of China
Yaohong Ma

Shandong Key Laboratory of Biosensor, Biology Institute of Shandong Academy of Sciences, Jinan, People’s Republic of China
Tengfei Wang

State Key Laboratory of Biobased Material and Green Papermaking (LBMP), Qilu University of Technology (Shandong Academy of Sciences), Jinan, People’s Republic of China

Download PDF

Sections

Plain Text BibTeX RIS

APA Style

Wenhu Chen, Runlong Ma, Ping Lin, Haibo Yuan, Hongling Liu, et al. (2022). Display of Glucose Oxidase on Bacillus subtilis Spore Surface and Its Application in Electrode. World Journal of Applied Chemistry, 7(2), 34-47. https://doi.org/10.11648/j.wjac.20220702.11

Copy | Download

ACS Style

Wenhu Chen; Runlong Ma; Ping Lin; Haibo Yuan; Hongling Liu, et al. Display of Glucose Oxidase on Bacillus subtilis Spore Surface and Its Application in Electrode. World J. Appl. Chem. 2022, 7(2), 34-47. doi: 10.11648/j.wjac.20220702.11

Copy | Download

AMA Style

Wenhu Chen, Runlong Ma, Ping Lin, Haibo Yuan, Hongling Liu, et al. Display of Glucose Oxidase on Bacillus subtilis Spore Surface and Its Application in Electrode. World J Appl Chem. 2022;7(2):34-47. doi: 10.11648/j.wjac.20220702.11

Copy | Download

@article{10.11648/j.wjac.20220702.11,
  author = {Wenhu Chen and Runlong Ma and Ping Lin and Haibo Yuan and Hongling Liu and Yi Jiang and Di Huang and Yaohong Ma and Tengfei Wang},
  title = {Display of Glucose Oxidase on Bacillus subtilis Spore Surface and Its Application in Electrode},
  journal = {World Journal of Applied Chemistry},
  volume = {7},
  number = {2},
  pages = {34-47},
  doi = {10.11648/j.wjac.20220702.11},
  url = {https://doi.org/10.11648/j.wjac.20220702.11},
  eprint = {https://article.sciencepublishinggroup.com/pdf/10.11648.j.wjac.20220702.11},
  abstract = {Glucose oxidase (GOD) is widely used in the fields of food, chemistry and medicine due to its specific catalytic activity, and it is the main tool enzyme in the biological field. The biological preparation of GOD can be realized by the method of surface display on Bacillus subtilis spores. In this study, we analyzed the display characteristics of eleven cot proteins on the spore coat and determined the fluorescence intensity of different spore coat proteins by flow cytometry as follows: CotX > CotY > CotF > CotC > CotE > CotG > CotV > CotB > CotM > CotA > CotW. Next, CotC, CotX and CotY were selected to display GOD on the surface of spores, and CotX demonstrated the highest GOD activity. Subsequently, graphene oxide was added to the glassy carbon electrode and prussian blue was deposited, the surface of this electrode was fixed with spore–GOD and finally covered with a layer of Nafion solution to prepare glucose detection electrode. In the range of 0.1-7.0 mmol/L glucose, the cyclic voltammetry curve on the enzyme electrode showed a linear relationship with the glucose concentration. The calibration curve equation was I = 1.3047Cglucose + 3.639 (R2 = 0.9929), and its detection limit was 7.5 μmol/L (S/N = 3). This modified electrode has good conductivity, stability, reproducibility and can be used for the analysis and determination of glucose.},
 year = {2022}
}

Copy | Download

TY  - JOUR
T1  - Display of Glucose Oxidase on Bacillus subtilis Spore Surface and Its Application in Electrode
AU  - Wenhu Chen
AU  - Runlong Ma
AU  - Ping Lin
AU  - Haibo Yuan
AU  - Hongling Liu
AU  - Yi Jiang
AU  - Di Huang
AU  - Yaohong Ma
AU  - Tengfei Wang
Y1  - 2022/04/08
PY  - 2022
N1  - https://doi.org/10.11648/j.wjac.20220702.11
DO  - 10.11648/j.wjac.20220702.11
T2  - World Journal of Applied Chemistry
JF  - World Journal of Applied Chemistry
JO  - World Journal of Applied Chemistry
SP  - 34
EP  - 47
PB  - Science Publishing Group
SN  - 2637-5982
UR  - https://doi.org/10.11648/j.wjac.20220702.11
AB  - Glucose oxidase (GOD) is widely used in the fields of food, chemistry and medicine due to its specific catalytic activity, and it is the main tool enzyme in the biological field. The biological preparation of GOD can be realized by the method of surface display on Bacillus subtilis spores. In this study, we analyzed the display characteristics of eleven cot proteins on the spore coat and determined the fluorescence intensity of different spore coat proteins by flow cytometry as follows: CotX > CotY > CotF > CotC > CotE > CotG > CotV > CotB > CotM > CotA > CotW. Next, CotC, CotX and CotY were selected to display GOD on the surface of spores, and CotX demonstrated the highest GOD activity. Subsequently, graphene oxide was added to the glassy carbon electrode and prussian blue was deposited, the surface of this electrode was fixed with spore–GOD and finally covered with a layer of Nafion solution to prepare glucose detection electrode. In the range of 0.1-7.0 mmol/L glucose, the cyclic voltammetry curve on the enzyme electrode showed a linear relationship with the glucose concentration. The calibration curve equation was I = 1.3047Cglucose + 3.639 (R2 = 0.9929), and its detection limit was 7.5 μmol/L (S/N = 3). This modified electrode has good conductivity, stability, reproducibility and can be used for the analysis and determination of glucose.
VL  - 7
IS  - 2
ER  -

Copy | Download