TY  - CONF
AU  - Pantić, Nevena
AU  - Spasojević, Milica
AU  - Prokopijević, Miloš
AU  - Spasojević, Dragica
AU  - Balaž, Ana Marija
AU  - Prodanović, Radivoje
AU  - Prodanović, Olivera
PY  - 2022
UR  - http://rimsi.imsi.bg.ac.rs/handle/123456789/1774
AB  - For the purpose of immobilization, one of the most commonly used enzymes is horseradish peroxidase (HRP). Different carriers can be used as supports for the immobilization of HRP: alginate, pectin, magnetic-beads, macroporous copolymers, silicas etc. Covalent binding of an enzyme to the carrier leads to the formation of strong linkage, thus preventing the enzyme leakage. Macroporous copolymers with different porous characteristics were used for the immobilization of horseradish peroxidase by employing periodate and glutaraldehyde method. Five and 25 mg of HRP were immobilized per gram of the copolymer. Increasing the amount of added enzyme leads to the increase of specific activity of immobilized enzyme. Copolymer with the pore diameter of 297 nm showed the most promising results in terms of specific activity. Immobilized enzymes can be used for the removal of phenolic compounds from waste effluents.
PB  - University of Belgrade, Technical Faculty in Bor
C3  - 29th International Conference Ecological Truth and Environmental Research
T1  - Covalent immobilization of horseradish peroxidase on novel macroporous poly(GMA-co-EGDMA) for phenol removal
EP  - 359
SP  - 354
UR  - https://hdl.handle.net/21.15107/rcub_rimsi_1774
ER  - 

@conference{
author = "Pantić, Nevena and Spasojević, Milica and Prokopijević, Miloš and Spasojević, Dragica and Balaž, Ana Marija and Prodanović, Radivoje and Prodanović, Olivera",
year = "2022",
abstract = "For the purpose of immobilization, one of the most commonly used enzymes is horseradish peroxidase (HRP). Different carriers can be used as supports for the immobilization of HRP: alginate, pectin, magnetic-beads, macroporous copolymers, silicas etc. Covalent binding of an enzyme to the carrier leads to the formation of strong linkage, thus preventing the enzyme leakage. Macroporous copolymers with different porous characteristics were used for the immobilization of horseradish peroxidase by employing periodate and glutaraldehyde method. Five and 25 mg of HRP were immobilized per gram of the copolymer. Increasing the amount of added enzyme leads to the increase of specific activity of immobilized enzyme. Copolymer with the pore diameter of 297 nm showed the most promising results in terms of specific activity. Immobilized enzymes can be used for the removal of phenolic compounds from waste effluents.",
publisher = "University of Belgrade, Technical Faculty in Bor",
journal = "29th International Conference Ecological Truth and Environmental Research",
title = "Covalent immobilization of horseradish peroxidase on novel macroporous poly(GMA-co-EGDMA) for phenol removal",
pages = "359-354",
url = "https://hdl.handle.net/21.15107/rcub_rimsi_1774"
}

Pantić, N., Spasojević, M., Prokopijević, M., Spasojević, D., Balaž, A. M., Prodanović, R.,& Prodanović, O.. (2022). Covalent immobilization of horseradish peroxidase on novel macroporous poly(GMA-co-EGDMA) for phenol removal. in 29th International Conference Ecological Truth and Environmental Research
University of Belgrade, Technical Faculty in Bor., 354-359.
https://hdl.handle.net/21.15107/rcub_rimsi_1774

Pantić N, Spasojević M, Prokopijević M, Spasojević D, Balaž AM, Prodanović R, Prodanović O. Covalent immobilization of horseradish peroxidase on novel macroporous poly(GMA-co-EGDMA) for phenol removal. in 29th International Conference Ecological Truth and Environmental Research. 2022;:354-359.
https://hdl.handle.net/21.15107/rcub_rimsi_1774 .

Pantić, Nevena, Spasojević, Milica, Prokopijević, Miloš, Spasojević, Dragica, Balaž, Ana Marija, Prodanović, Radivoje, Prodanović, Olivera, "Covalent immobilization of horseradish peroxidase on novel macroporous poly(GMA-co-EGDMA) for phenol removal" in 29th International Conference Ecological Truth and Environmental Research (2022):354-359,
https://hdl.handle.net/21.15107/rcub_rimsi_1774 .

TY  - JOUR
AU  - Kovačević, G.
AU  - Elgahwash, R.G.A.
AU  - Blažić, M.
AU  - Pantić, Nevena
AU  - Prodanović, Olivera
AU  - Balaž, Ana Marija
AU  - Prodanović, Radivoje
PY  - 2022
UR  - http://rimsi.imsi.bg.ac.rs/handle/123456789/1543
AB  - Saccharomyces cerevisiae, known as bakers’ yeast, is one of the most utilized yeasts in industry. Several enzymes that are naturally produced by yeast, such as invertase and catalase, combined with heterologously expressed glucose oxidase (GOx), represent the enzyme machinery for fructose and gluconic acid production. Therefore, we have used yeast cell walls with expressed glucose oxidase as a platform for crosslinking with invertase and catalase to create biocatalyst cells for the high yield sucrose conversion. Using 5% (w/v) suspension of cross-linked yeast cell walls in 0.15 M sucrose solution, 1.86 g L−1 h−1 of gluconic acid has been obtained using wt-GOx, while mutant A2-GOx produced 2.91 g L−1 h−1 of gluconic acid. Increasing the concentration of modified yeast cells walls to 10% (w/v) we were able to obtain almost 100% conversion of glucose to gluconic acid using A2-GOx in the first cycle. Reusing the modified cells walls in three consecutive cycles, conversion dropped to approximately 70% using A2-GOx and 40% using wt-GOx.
PB  - Elsevier B.V.
T2  - Molecular Catalysis
T1  - Production of fructose and gluconic acid from sucrose with cross-linked yeast cell walls expressing glucose oxidase on the surface
VL  - 522
DO  - 10.1016/j.mcat.2022.112215
ER  - 

@article{
author = "Kovačević, G. and Elgahwash, R.G.A. and Blažić, M. and Pantić, Nevena and Prodanović, Olivera and Balaž, Ana Marija and Prodanović, Radivoje",
year = "2022",
abstract = "Saccharomyces cerevisiae, known as bakers’ yeast, is one of the most utilized yeasts in industry. Several enzymes that are naturally produced by yeast, such as invertase and catalase, combined with heterologously expressed glucose oxidase (GOx), represent the enzyme machinery for fructose and gluconic acid production. Therefore, we have used yeast cell walls with expressed glucose oxidase as a platform for crosslinking with invertase and catalase to create biocatalyst cells for the high yield sucrose conversion. Using 5% (w/v) suspension of cross-linked yeast cell walls in 0.15 M sucrose solution, 1.86 g L−1 h−1 of gluconic acid has been obtained using wt-GOx, while mutant A2-GOx produced 2.91 g L−1 h−1 of gluconic acid. Increasing the concentration of modified yeast cells walls to 10% (w/v) we were able to obtain almost 100% conversion of glucose to gluconic acid using A2-GOx in the first cycle. Reusing the modified cells walls in three consecutive cycles, conversion dropped to approximately 70% using A2-GOx and 40% using wt-GOx.",
publisher = "Elsevier B.V.",
journal = "Molecular Catalysis",
title = "Production of fructose and gluconic acid from sucrose with cross-linked yeast cell walls expressing glucose oxidase on the surface",
volume = "522",
doi = "10.1016/j.mcat.2022.112215"
}

Kovačević, G., Elgahwash, R.G.A., Blažić, M., Pantić, N., Prodanović, O., Balaž, A. M.,& Prodanović, R.. (2022). Production of fructose and gluconic acid from sucrose with cross-linked yeast cell walls expressing glucose oxidase on the surface. in Molecular Catalysis
Elsevier B.V.., 522.
https://doi.org/10.1016/j.mcat.2022.112215

Kovačević G, Elgahwash R, Blažić M, Pantić N, Prodanović O, Balaž AM, Prodanović R. Production of fructose and gluconic acid from sucrose with cross-linked yeast cell walls expressing glucose oxidase on the surface. in Molecular Catalysis. 2022;522.
doi:10.1016/j.mcat.2022.112215 .

Kovačević, G., Elgahwash, R.G.A., Blažić, M., Pantić, Nevena, Prodanović, Olivera, Balaž, Ana Marija, Prodanović, Radivoje, "Production of fructose and gluconic acid from sucrose with cross-linked yeast cell walls expressing glucose oxidase on the surface" in Molecular Catalysis, 522 (2022),
https://doi.org/10.1016/j.mcat.2022.112215 . .

TY  - JOUR
AU  - Pantić, Nevena
AU  - Spasojević, Milica
AU  - Stojanović, Zeljko P
AU  - Veljović, Đorđe
AU  - Krstic, Jugoslav
AU  - Balaž, Ana Marija
AU  - Prodanović, Radivoje
AU  - Prodanović, Olivera
PY  - 2022
UR  - http://rimsi.imsi.bg.ac.rs/handle/123456789/1530
AB  - Novel macroporous copolymers of glycidyl methacrylate and ethylene glycol dimethacrylate with mean pore size diameters ranging from 150 to 310 nm were synthesized by dispersion polymerization and modified with ethylenediamine. The glutaraldehyde and periodate method were employed to immobilize horseradish peroxidase (HRP) onto these carriers. The activity of the immobilized enzyme was greatly affected by the pore size of the carrier. The highest specific activities of 9.65 and 8.94 U/g of dry weight were obtained for HRP immobilized by the periodate-route onto poly(GMA-co-EGDMA) carriers with pore size diameters of 234 and 297 nm, respectively. Stability studies showed an improved operational stability of immobilized peroxidase at 65 degrees C and in an organic solvent. HRP immobilized on a copolymer with a pore size of 234 nm, showing the highest specific activity and good stability, had higher activities at almost all pH values than the native enzyme and the increased K-m value for pyrogallol oxidation. Immobilized HRP retained 80% of its original activity after five consecutive cycles of the pyrogallol oxidation and 98% of its initial activity in a storage stability study. Enzyme immobilized onto the macroporous copolymer with the pore size diameter of 234 nm showed a substantial degree of phenol removal achieved by immobilized peroxidase.
PB  - Springer, New York
T2  - Journal of Polymers and the Environment
T1  - Immobilization of Horseradish Peroxidase on Macroporous Glycidyl-Based Copolymers with Different Surface Characteristics for the Removal of Phenol
DO  - 10.1007/s10924-021-02364-3
ER  - 

@article{
author = "Pantić, Nevena and Spasojević, Milica and Stojanović, Zeljko P and Veljović, Đorđe and Krstic, Jugoslav and Balaž, Ana Marija and Prodanović, Radivoje and Prodanović, Olivera",
year = "2022",
abstract = "Novel macroporous copolymers of glycidyl methacrylate and ethylene glycol dimethacrylate with mean pore size diameters ranging from 150 to 310 nm were synthesized by dispersion polymerization and modified with ethylenediamine. The glutaraldehyde and periodate method were employed to immobilize horseradish peroxidase (HRP) onto these carriers. The activity of the immobilized enzyme was greatly affected by the pore size of the carrier. The highest specific activities of 9.65 and 8.94 U/g of dry weight were obtained for HRP immobilized by the periodate-route onto poly(GMA-co-EGDMA) carriers with pore size diameters of 234 and 297 nm, respectively. Stability studies showed an improved operational stability of immobilized peroxidase at 65 degrees C and in an organic solvent. HRP immobilized on a copolymer with a pore size of 234 nm, showing the highest specific activity and good stability, had higher activities at almost all pH values than the native enzyme and the increased K-m value for pyrogallol oxidation. Immobilized HRP retained 80% of its original activity after five consecutive cycles of the pyrogallol oxidation and 98% of its initial activity in a storage stability study. Enzyme immobilized onto the macroporous copolymer with the pore size diameter of 234 nm showed a substantial degree of phenol removal achieved by immobilized peroxidase.",
publisher = "Springer, New York",
journal = "Journal of Polymers and the Environment",
title = "Immobilization of Horseradish Peroxidase on Macroporous Glycidyl-Based Copolymers with Different Surface Characteristics for the Removal of Phenol",
doi = "10.1007/s10924-021-02364-3"
}

Pantić, N., Spasojević, M., Stojanović, Z. P., Veljović, Đ., Krstic, J., Balaž, A. M., Prodanović, R.,& Prodanović, O.. (2022). Immobilization of Horseradish Peroxidase on Macroporous Glycidyl-Based Copolymers with Different Surface Characteristics for the Removal of Phenol. in Journal of Polymers and the Environment
Springer, New York..
https://doi.org/10.1007/s10924-021-02364-3

Pantić N, Spasojević M, Stojanović ZP, Veljović Đ, Krstic J, Balaž AM, Prodanović R, Prodanović O. Immobilization of Horseradish Peroxidase on Macroporous Glycidyl-Based Copolymers with Different Surface Characteristics for the Removal of Phenol. in Journal of Polymers and the Environment. 2022;.
doi:10.1007/s10924-021-02364-3 .

Pantić, Nevena, Spasojević, Milica, Stojanović, Zeljko P, Veljović, Đorđe, Krstic, Jugoslav, Balaž, Ana Marija, Prodanović, Radivoje, Prodanović, Olivera, "Immobilization of Horseradish Peroxidase on Macroporous Glycidyl-Based Copolymers with Different Surface Characteristics for the Removal of Phenol" in Journal of Polymers and the Environment (2022),
https://doi.org/10.1007/s10924-021-02364-3 . .

TY  - JOUR
AU  - Balaž, Ana Marija
AU  - BLazic, Marija B.
AU  - Popović, Nikolina
AU  - Prodanović, Olivera
AU  - Ostafe, Raluca
AU  - Fischer, Rainer
AU  - Prodanović, Radivoje
PY  - 2020
UR  - http://rimsi.imsi.bg.ac.rs/handle/123456789/1297
AB  - Production of soluble cellobiose dehydrogenase (CDH) mutant proteins previously evolved on the surface of S. cerevisiae yeast cells was established for use in biosensors and biofuel cells. For this purpose, mutant cdh genes tm (D20N, A64T, V592M), H5 (D20N, V22A, A64T, V592M) and H9 (D20N, A64T, T84A, A261P, V592M, E674G, N715S) were cloned to pPICZ alpha plasmid and transformed into Pichia pastoris KM71H strain for high expression in a soluble form and kinetic characterization. After 6 days of expression under methanol induction, the CDHs were purified by ultrafiltration, ion-exchange chromatography and gel filtration. Sodium dodecyl sulfate electrophoresis confirmed the purity and presence of a single protein band at a molecular weight of 100 kDa. Kinetic characterization showed that the H5 mutant had the highest catalytic constant of 43.5 s(-1) for lactose, while the mutant H9 showed the highest specificity constant for lactose of 132 mM(-1) s(-1). All three mutant proteins did not change the pH optimum that was between 4.5 and 5.5. Compared to the previously obtained wild types and mutants of CDH from Phanerochaete chrysosporium, the variants reported in this article had higher activity and specificity that together with high protein expression rate in P. pastoris, makes them good candidates for use in biotechnology for lactobionic acid production and biosensor manufacture.
PB  - Srpsko hemijsko društvo, Beograd
T2  - Journal of the Serbian Chemical Society
T1  - Expression, purification and characterization of cellobiose dehydrogenase mutants from Phanerochaete chrysosporium in Pichia pastoris KM71H strain
EP  - 35
IS  - 1
SP  - 25
VL  - 85
DO  - 10.2298/JSC190320058B
ER  - 

@article{
author = "Balaž, Ana Marija and BLazic, Marija B. and Popović, Nikolina and Prodanović, Olivera and Ostafe, Raluca and Fischer, Rainer and Prodanović, Radivoje",
year = "2020",
abstract = "Production of soluble cellobiose dehydrogenase (CDH) mutant proteins previously evolved on the surface of S. cerevisiae yeast cells was established for use in biosensors and biofuel cells. For this purpose, mutant cdh genes tm (D20N, A64T, V592M), H5 (D20N, V22A, A64T, V592M) and H9 (D20N, A64T, T84A, A261P, V592M, E674G, N715S) were cloned to pPICZ alpha plasmid and transformed into Pichia pastoris KM71H strain for high expression in a soluble form and kinetic characterization. After 6 days of expression under methanol induction, the CDHs were purified by ultrafiltration, ion-exchange chromatography and gel filtration. Sodium dodecyl sulfate electrophoresis confirmed the purity and presence of a single protein band at a molecular weight of 100 kDa. Kinetic characterization showed that the H5 mutant had the highest catalytic constant of 43.5 s(-1) for lactose, while the mutant H9 showed the highest specificity constant for lactose of 132 mM(-1) s(-1). All three mutant proteins did not change the pH optimum that was between 4.5 and 5.5. Compared to the previously obtained wild types and mutants of CDH from Phanerochaete chrysosporium, the variants reported in this article had higher activity and specificity that together with high protein expression rate in P. pastoris, makes them good candidates for use in biotechnology for lactobionic acid production and biosensor manufacture.",
publisher = "Srpsko hemijsko društvo, Beograd",
journal = "Journal of the Serbian Chemical Society",
title = "Expression, purification and characterization of cellobiose dehydrogenase mutants from Phanerochaete chrysosporium in Pichia pastoris KM71H strain",
pages = "35-25",
number = "1",
volume = "85",
doi = "10.2298/JSC190320058B"
}

Balaž, A. M., BLazic, M. B., Popović, N., Prodanović, O., Ostafe, R., Fischer, R.,& Prodanović, R.. (2020). Expression, purification and characterization of cellobiose dehydrogenase mutants from Phanerochaete chrysosporium in Pichia pastoris KM71H strain. in Journal of the Serbian Chemical Society
Srpsko hemijsko društvo, Beograd., 85(1), 25-35.
https://doi.org/10.2298/JSC190320058B

Balaž AM, BLazic MB, Popović N, Prodanović O, Ostafe R, Fischer R, Prodanović R. Expression, purification and characterization of cellobiose dehydrogenase mutants from Phanerochaete chrysosporium in Pichia pastoris KM71H strain. in Journal of the Serbian Chemical Society. 2020;85(1):25-35.
doi:10.2298/JSC190320058B .

Balaž, Ana Marija, BLazic, Marija B., Popović, Nikolina, Prodanović, Olivera, Ostafe, Raluca, Fischer, Rainer, Prodanović, Radivoje, "Expression, purification and characterization of cellobiose dehydrogenase mutants from Phanerochaete chrysosporium in Pichia pastoris KM71H strain" in Journal of the Serbian Chemical Society, 85, no. 1 (2020):25-35,
https://doi.org/10.2298/JSC190320058B . .

TY  - JOUR
AU  - Spasojević, Milica
AU  - Prodanović, Olivera
AU  - Pantić, Nevena
AU  - Popović, Nikolina
AU  - Balaž, Ana Marija
AU  - Prodanović, Radivoje
PY  - 2019
UR  - http://rimsi.imsi.bg.ac.rs/handle/123456789/1757
AB  - Strategies based on the enzyme application are increasingly replacing the conventional chemical procedures because of their efficiency, quicker performance and environmental protection. However, natural enzymes can rarely be used in industry since their beneficial features cannot endure the industrial conditions. Additional drawbacks of natural enzymes are their inhibition by reaction products and difficulty to be removed from the reaction mixture. The most promising technique to substantially improve the enzyme properties, such as activity, pH, thermal and organic-solvent stability, reusability and storage stability, in non-natural environments is by the enzyme immobilization. In this review different techniques used to immobilize enzymes to inert carriers were summarized. Different materials of both the organic and inorganic origin were used as carriers for the enzyme immobilization. A class of new materials where the enzyme performance was enhanced by combining different classical materials and shaping in specific forms was also summarized.
PB  - Faculty of Technology Zvornik
T2  - Journal of Engineering & Processing Management
T1  - The enzyme immobilization: Carriers and immobilization methods
EP  - 105
IS  - 2
SP  - 89
VL  - 11
DO  - 10.7251/jepm1902089s
ER  - 

@article{
author = "Spasojević, Milica and Prodanović, Olivera and Pantić, Nevena and Popović, Nikolina and Balaž, Ana Marija and Prodanović, Radivoje",
year = "2019",
abstract = "Strategies based on the enzyme application are increasingly replacing the conventional chemical procedures because of their efficiency, quicker performance and environmental protection. However, natural enzymes can rarely be used in industry since their beneficial features cannot endure the industrial conditions. Additional drawbacks of natural enzymes are their inhibition by reaction products and difficulty to be removed from the reaction mixture. The most promising technique to substantially improve the enzyme properties, such as activity, pH, thermal and organic-solvent stability, reusability and storage stability, in non-natural environments is by the enzyme immobilization. In this review different techniques used to immobilize enzymes to inert carriers were summarized. Different materials of both the organic and inorganic origin were used as carriers for the enzyme immobilization. A class of new materials where the enzyme performance was enhanced by combining different classical materials and shaping in specific forms was also summarized.",
publisher = "Faculty of Technology Zvornik",
journal = "Journal of Engineering & Processing Management",
title = "The enzyme immobilization: Carriers and immobilization methods",
pages = "105-89",
number = "2",
volume = "11",
doi = "10.7251/jepm1902089s"
}

Spasojević, M., Prodanović, O., Pantić, N., Popović, N., Balaž, A. M.,& Prodanović, R.. (2019). The enzyme immobilization: Carriers and immobilization methods. in Journal of Engineering & Processing Management
Faculty of Technology Zvornik., 11(2), 89-105.
https://doi.org/10.7251/jepm1902089s

Spasojević M, Prodanović O, Pantić N, Popović N, Balaž AM, Prodanović R. The enzyme immobilization: Carriers and immobilization methods. in Journal of Engineering & Processing Management. 2019;11(2):89-105.
doi:10.7251/jepm1902089s .

Spasojević, Milica, Prodanović, Olivera, Pantić, Nevena, Popović, Nikolina, Balaž, Ana Marija, Prodanović, Radivoje, "The enzyme immobilization: Carriers and immobilization methods" in Journal of Engineering & Processing Management, 11, no. 2 (2019):89-105,
https://doi.org/10.7251/jepm1902089s . .

TY  - JOUR
AU  - Blazic, Marija
AU  - Balaž, Ana Marija
AU  - Prodanović, Olivera
AU  - Popović, Nikolina
AU  - Ostafe, Raluca
AU  - Fischer, Rainer
AU  - Prodanović, Radivoje
PY  - 2019
UR  - http://rimsi.imsi.bg.ac.rs/handle/123456789/1240
AB  - Featured Application Developed fluorescent assay and expression system can be used for obtaining improved cellobiose dehydrogenase whole cell biocatalysts for lactobionic acid production and building of biosensors and biofuel cells. Cellobiose dehydrogenase (CDH) from Phanerochaete chrysosporium can be used in lactobionic acid production, biosensor for lactose, biofuel cells, lignocellulose degradation, and wound-healing applications. To make it a better biocatalyst, CDH with higher activity in an immobilized form is desirable. For this purpose, CDH was expressed for the first time on the surface of S. cerevisiae EBY100 cells in an active form as a triple mutant tmCDH (D20N, A64T, V592M) and evolved further for higher activity using resazurin-based fluorescent assay. In order to decrease blank reaction of resazurin with yeast cells and to have linear correlation between enzyme activity on the cell surface and fluorescence signal, the assay was optimized with respect to resazurin concentration (0.1 mM), substrate concentration (10 mM lactose and 0.08 mM cellobiose), and pH (6.0). Using optimized assay an error prone PCR gene library of tmCDH was screened. Two mutants with 5 (H5) and 7 mutations (H9) were found having two times higher activity than the parent tmCDH enzyme that already had improved activity compared to wild type CDH whose activity could not be detected on the surface of yeast cells.
PB  - MDPI, Basel
T2  - Applied Sciences-Basel
T1  - Directed Evolution of Cellobiose Dehydrogenase on the Surface of Yeast Cells Using Resazurin-Based Fluorescent Assay
IS  - 7
VL  - 9
DO  - 10.3390/app9071413
ER  - 

@article{
author = "Blazic, Marija and Balaž, Ana Marija and Prodanović, Olivera and Popović, Nikolina and Ostafe, Raluca and Fischer, Rainer and Prodanović, Radivoje",
year = "2019",
abstract = "Featured Application Developed fluorescent assay and expression system can be used for obtaining improved cellobiose dehydrogenase whole cell biocatalysts for lactobionic acid production and building of biosensors and biofuel cells. Cellobiose dehydrogenase (CDH) from Phanerochaete chrysosporium can be used in lactobionic acid production, biosensor for lactose, biofuel cells, lignocellulose degradation, and wound-healing applications. To make it a better biocatalyst, CDH with higher activity in an immobilized form is desirable. For this purpose, CDH was expressed for the first time on the surface of S. cerevisiae EBY100 cells in an active form as a triple mutant tmCDH (D20N, A64T, V592M) and evolved further for higher activity using resazurin-based fluorescent assay. In order to decrease blank reaction of resazurin with yeast cells and to have linear correlation between enzyme activity on the cell surface and fluorescence signal, the assay was optimized with respect to resazurin concentration (0.1 mM), substrate concentration (10 mM lactose and 0.08 mM cellobiose), and pH (6.0). Using optimized assay an error prone PCR gene library of tmCDH was screened. Two mutants with 5 (H5) and 7 mutations (H9) were found having two times higher activity than the parent tmCDH enzyme that already had improved activity compared to wild type CDH whose activity could not be detected on the surface of yeast cells.",
publisher = "MDPI, Basel",
journal = "Applied Sciences-Basel",
title = "Directed Evolution of Cellobiose Dehydrogenase on the Surface of Yeast Cells Using Resazurin-Based Fluorescent Assay",
number = "7",
volume = "9",
doi = "10.3390/app9071413"
}

Blazic, M., Balaž, A. M., Prodanović, O., Popović, N., Ostafe, R., Fischer, R.,& Prodanović, R.. (2019). Directed Evolution of Cellobiose Dehydrogenase on the Surface of Yeast Cells Using Resazurin-Based Fluorescent Assay. in Applied Sciences-Basel
MDPI, Basel., 9(7).
https://doi.org/10.3390/app9071413

Blazic M, Balaž AM, Prodanović O, Popović N, Ostafe R, Fischer R, Prodanović R. Directed Evolution of Cellobiose Dehydrogenase on the Surface of Yeast Cells Using Resazurin-Based Fluorescent Assay. in Applied Sciences-Basel. 2019;9(7).
doi:10.3390/app9071413 .

Blazic, Marija, Balaž, Ana Marija, Prodanović, Olivera, Popović, Nikolina, Ostafe, Raluca, Fischer, Rainer, Prodanović, Radivoje, "Directed Evolution of Cellobiose Dehydrogenase on the Surface of Yeast Cells Using Resazurin-Based Fluorescent Assay" in Applied Sciences-Basel, 9, no. 7 (2019),
https://doi.org/10.3390/app9071413 . .