Blažić, Marija

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  • Blažić, Marija (13)
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Expression, purification and characterization of cellobiose dehydrogenase mutants from Phanerochaete chrysosporium in Pichia pastoris KM71H strain

Balaž, Ana Marija; Blažić, Marija; Popović, Nikolina; Prodanović, Olivera; Ostafe, Raluca; Fischer, Rainer; Prodanović, Radivoje

(Belgrade : Serbian Chemical Society, 2020)

TY  - JOUR
AU  - Balaž, Ana Marija
AU  - Blažić, Marija
AU  - Popović, Nikolina
AU  - Prodanović, Olivera
AU  - Ostafe, Raluca
AU  - Fischer, Rainer
AU  - Prodanović, Radivoje
PY  - 2020
UR  - http://cherry.chem.bg.ac.rs/handle/123456789/4270
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 pPICZα 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.
AB  - У циљу употребе у биосензорима и биогоривним ћелијама, успостављена је производњарастворних облика целобиоза дехидрогеназе (CDH) претходно еволуираних на површиниквашчевих ћелија S. cerevisiae. У ту сврху су мутанти CDH, tm (D20N, A64T, V592M), H5(D20N, V22A, A64T, V592M) и H9 (D20N, A64T, T84A, A261P, V592M, E674G, N715S)клонирани у pPICZα плазмид и трансформисани у Pichia pastoris KM71H сој за високуекспресију у растворном облику и кинетичку карактеризацију. После 6 дана експресије подиндукцијом метанолом, мутанти су пречишћени ултрафилтрацијом, јоноизмењивачкомхроматографијом и гел-филтрацијом. SDS електрофореза је потврдила чистоћу уз присуствоједне протеинске траке молекулскe масe од 100 kDa. Кинетичка карактеризација је показалада H5 мутирани протеин поседује највећу каталитичку константу од 43,5 s-1 за лактозу, докје H9 имао највећу константу специфичности за лактозу од 132 mM-1 s-1. Сва три мутиранапротеина су имала неизмењен pH оптимум који је био у опсегу од 4,5 до 5,5. У поређењу сапретходно добијеним природним и мутантним облицима CDH протеина из Phanerochaetechrysosporium, облици приказани у овом раду имају већу активност и специфичност, што их,повезано са високом експресијом протеина у P. Pastoris, чини добрим кандидатима за упо-требу у биотехнологији за производњу лактобионске киселине и биосензора.
PB  - Belgrade : Serbian Chemical Society
T2  - Journal of the Serbian Chemical Society
T1  - Expression, purification and characterization of cellobiose dehydrogenase mutants from Phanerochaete chrysosporium in Pichia pastoris KM71H strain
T1  - Ekspresija, prečišćavanje i karakterizacija mutanata celobioza - dehidrogenaze iz Phanerochaete chrysosporium u Pichia pastoris KM71H soju
VL  - 85
IS  - 1
SP  - 25
EP  - 35
DO  - 10.2298/JSC190320058B
ER  - 
@article{
author = "Balaž, Ana Marija and Blažić, Marija and Popović, Nikolina and Prodanović, Olivera and Ostafe, Raluca and Fischer, Rainer and Prodanović, Radivoje",
year = "2020",
url = "http://cherry.chem.bg.ac.rs/handle/123456789/4270",
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 pPICZα 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., У циљу употребе у биосензорима и биогоривним ћелијама, успостављена је производњарастворних облика целобиоза дехидрогеназе (CDH) претходно еволуираних на површиниквашчевих ћелија S. cerevisiae. У ту сврху су мутанти CDH, tm (D20N, A64T, V592M), H5(D20N, V22A, A64T, V592M) и H9 (D20N, A64T, T84A, A261P, V592M, E674G, N715S)клонирани у pPICZα плазмид и трансформисани у Pichia pastoris KM71H сој за високуекспресију у растворном облику и кинетичку карактеризацију. После 6 дана експресије подиндукцијом метанолом, мутанти су пречишћени ултрафилтрацијом, јоноизмењивачкомхроматографијом и гел-филтрацијом. SDS електрофореза је потврдила чистоћу уз присуствоједне протеинске траке молекулскe масe од 100 kDa. Кинетичка карактеризација је показалада H5 мутирани протеин поседује највећу каталитичку константу од 43,5 s-1 за лактозу, докје H9 имао највећу константу специфичности за лактозу од 132 mM-1 s-1. Сва три мутиранапротеина су имала неизмењен pH оптимум који је био у опсегу од 4,5 до 5,5. У поређењу сапретходно добијеним природним и мутантним облицима CDH протеина из Phanerochaetechrysosporium, облици приказани у овом раду имају већу активност и специфичност, што их,повезано са високом експресијом протеина у P. Pastoris, чини добрим кандидатима за упо-требу у биотехнологији за производњу лактобионске киселине и биосензора.",
publisher = "Belgrade : Serbian Chemical Society",
journal = "Journal of the Serbian Chemical Society",
title = "Expression, purification and characterization of cellobiose dehydrogenase mutants from Phanerochaete chrysosporium in Pichia pastoris KM71H strain, Ekspresija, prečišćavanje i karakterizacija mutanata celobioza - dehidrogenaze iz Phanerochaete chrysosporium u Pichia pastoris KM71H soju",
volume = "85",
number = "1",
pages = "25-35",
doi = "10.2298/JSC190320058B"
}
Balaž, A. M., Blažić, M., Popović, N., Prodanović, O., Ostafe, R., Fischer, R.,& Prodanović, R. (2020). Ekspresija, prečišćavanje i karakterizacija mutanata celobioza - dehidrogenaze iz Phanerochaete chrysosporium u Pichia pastoris KM71H soju.
Journal of the Serbian Chemical Society
Belgrade : Serbian Chemical Society., 85(1), 25-35.
https://doi.org/10.2298/JSC190320058B
Balaž AM, Blažić M, Popović N, Prodanović O, Ostafe R, Fischer R, Prodanović R. Ekspresija, prečišćavanje i karakterizacija mutanata celobioza - dehidrogenaze iz Phanerochaete chrysosporium u Pichia pastoris KM71H soju. Journal of the Serbian Chemical Society. 2020;85(1):25-35
Balaž Ana Marija, Blažić Marija, Popović Nikolina, Prodanović Olivera, Ostafe Raluca, Fischer Rainer, Prodanović Radivoje, "Ekspresija, prečišćavanje i karakterizacija mutanata celobioza - dehidrogenaze iz Phanerochaete chrysosporium u Pichia pastoris KM71H soju" Journal of the Serbian Chemical Society, 85, no. 1 (2020):25-35,
https://doi.org/10.2298/JSC190320058B .

Protein engineering of cellobiose dehydrogenase from Phanerochaete chrysosporium in yeast Saccharomyces cerevisiae InvSc1 for increased activity and stability

Blažić, Marija; Balaž, Ana Marija; Tadić, Vojin; Draganić, Bojana; Ostafe, Raluca; Fischer, Rainer; Prodanović, Radivoje

(Elsevier, 2019)

TY  - JOUR
AU  - Blažić, Marija
AU  - Balaž, Ana Marija
AU  - Tadić, Vojin
AU  - Draganić, Bojana
AU  - Ostafe, Raluca
AU  - Fischer, Rainer
AU  - Prodanović, Radivoje
PY  - 2019
UR  - http://cherry.chem.bg.ac.rs/handle/123456789/2898
AB  - Cellobiose dehydrogenase (CDH) can be used in industry for lactobionic acid production, as a part of biosensors for disaccharides and in wound healing. In fungi it is involved in lignocellulose degradation. CDH gene from Phanerochaete chrysosporium has been cloned in pYES2 plasmid for extracellular expression and protein engineering in yeast Saccharomyces cerevisiae InvSC1 for the first time. A CDH gene library was generated using error-prone PCR and screened by spectrophotometric enzymatic assay based on 2,6-dichloroindophenol reduction detection in microtiter plates. Several mutants with increased activity and specificity towards lactose and cellobiose were found, purified and characterized in detail. Recombinant CDH enzymes showed a broad molecular weight between 120 and 150 KDa due to hyper-glycosylation and the best S137 N mutant showed 2.2 times increased k cat and 1.5 and 2 times increased specificity constant for lactose and cellobiose compared to the wild type enzyme. pH optimum of mutants was not changed while thermostability of selected mutants improved and S137 N mutant retained 30% of it's original activity after 15 min at 70 °C compared to 10% of activity that the wild type enzyme retained. Mutants M65S and S137 N showed also 1.6 and 1.5 times increased productivity of hydrogen peroxide in the presence of 30 mM lactose compared to the wild type.
PB  - Elsevier
T2  - Biochemical Engineering Journal
T1  - Protein engineering of cellobiose dehydrogenase from Phanerochaete chrysosporium in yeast Saccharomyces cerevisiae InvSc1 for increased activity and stability
VL  - 146
SP  - 179
EP  - 185
DO  - 10.1016/j.bej.2019.03.025
ER  - 
@article{
author = "Blažić, Marija and Balaž, Ana Marija and Tadić, Vojin and Draganić, Bojana and Ostafe, Raluca and Fischer, Rainer and Prodanović, Radivoje",
year = "2019",
url = "http://cherry.chem.bg.ac.rs/handle/123456789/2898",
abstract = "Cellobiose dehydrogenase (CDH) can be used in industry for lactobionic acid production, as a part of biosensors for disaccharides and in wound healing. In fungi it is involved in lignocellulose degradation. CDH gene from Phanerochaete chrysosporium has been cloned in pYES2 plasmid for extracellular expression and protein engineering in yeast Saccharomyces cerevisiae InvSC1 for the first time. A CDH gene library was generated using error-prone PCR and screened by spectrophotometric enzymatic assay based on 2,6-dichloroindophenol reduction detection in microtiter plates. Several mutants with increased activity and specificity towards lactose and cellobiose were found, purified and characterized in detail. Recombinant CDH enzymes showed a broad molecular weight between 120 and 150 KDa due to hyper-glycosylation and the best S137 N mutant showed 2.2 times increased k cat and 1.5 and 2 times increased specificity constant for lactose and cellobiose compared to the wild type enzyme. pH optimum of mutants was not changed while thermostability of selected mutants improved and S137 N mutant retained 30% of it's original activity after 15 min at 70 °C compared to 10% of activity that the wild type enzyme retained. Mutants M65S and S137 N showed also 1.6 and 1.5 times increased productivity of hydrogen peroxide in the presence of 30 mM lactose compared to the wild type.",
publisher = "Elsevier",
journal = "Biochemical Engineering Journal",
title = "Protein engineering of cellobiose dehydrogenase from Phanerochaete chrysosporium in yeast Saccharomyces cerevisiae InvSc1 for increased activity and stability",
volume = "146",
pages = "179-185",
doi = "10.1016/j.bej.2019.03.025"
}
Blažić, M., Balaž, A. M., Tadić, V., Draganić, B., Ostafe, R., Fischer, R.,& Prodanović, R. (2019). Protein engineering of cellobiose dehydrogenase from Phanerochaete chrysosporium in yeast Saccharomyces cerevisiae InvSc1 for increased activity and stability.
Biochemical Engineering Journal
Elsevier., 146, 179-185.
https://doi.org/10.1016/j.bej.2019.03.025
Blažić M, Balaž AM, Tadić V, Draganić B, Ostafe R, Fischer R, Prodanović R. Protein engineering of cellobiose dehydrogenase from Phanerochaete chrysosporium in yeast Saccharomyces cerevisiae InvSc1 for increased activity and stability. Biochemical Engineering Journal. 2019;146:179-185
Blažić Marija, Balaž Ana Marija, Tadić Vojin, Draganić Bojana, Ostafe Raluca, Fischer Rainer, Prodanović Radivoje, "Protein engineering of cellobiose dehydrogenase from Phanerochaete chrysosporium in yeast Saccharomyces cerevisiae InvSc1 for increased activity and stability" Biochemical Engineering Journal, 146 (2019):179-185,
https://doi.org/10.1016/j.bej.2019.03.025 .
6
1
6

Supplementary data for the article: Kostić, A. Ž.; Gašić, U. M.; Pešić, M. B.; Stanojević, S. P.; Barać, M. B.; Mačukanović-Jocić, M. P.; Avramov, S. N.; Tešić, Ž. L. Phytochemical Analysis and Total Antioxidant Capacity of Rhizome, Above-Ground Vegetative Parts and Flower of Three Iris Species. Chemistry and Biodiversity 2019, 16 (3), 1–17. https://doi.org/10.1002/cbdv.201800565

Blažić, Marija; Balaž, Ana Marija; Tadić, Vojin; Draganić, Bojana; Ostafe, Raluca; Fischer, Rainer; Prodanović, Radivoje

(Elsevier, 2019)

TY  - BOOK
AU  - Blažić, Marija
AU  - Balaž, Ana Marija
AU  - Tadić, Vojin
AU  - Draganić, Bojana
AU  - Ostafe, Raluca
AU  - Fischer, Rainer
AU  - Prodanović, Radivoje
PY  - 2019
UR  - http://cherry.chem.bg.ac.rs/handle/123456789/2900
PB  - Elsevier
T2  - Biochemical Engineering Journal
T1  - Supplementary data for the article: Kostić, A. Ž.; Gašić, U. M.; Pešić, M. B.; Stanojević, S. P.; Barać, M. B.; Mačukanović-Jocić, M. P.; Avramov, S. N.; Tešić, Ž. L. Phytochemical Analysis and Total Antioxidant Capacity of Rhizome, Above-Ground Vegetative Parts and Flower of Three Iris Species. Chemistry and Biodiversity 2019, 16 (3), 1–17. https://doi.org/10.1002/cbdv.201800565
ER  - 
@book{
author = "Blažić, Marija and Balaž, Ana Marija and Tadić, Vojin and Draganić, Bojana and Ostafe, Raluca and Fischer, Rainer and Prodanović, Radivoje",
year = "2019",
url = "http://cherry.chem.bg.ac.rs/handle/123456789/2900",
publisher = "Elsevier",
journal = "Biochemical Engineering Journal",
title = "Supplementary data for the article: Kostić, A. Ž.; Gašić, U. M.; Pešić, M. B.; Stanojević, S. P.; Barać, M. B.; Mačukanović-Jocić, M. P.; Avramov, S. N.; Tešić, Ž. L. Phytochemical Analysis and Total Antioxidant Capacity of Rhizome, Above-Ground Vegetative Parts and Flower of Three Iris Species. Chemistry and Biodiversity 2019, 16 (3), 1–17. https://doi.org/10.1002/cbdv.201800565"
}
Blažić, M., Balaž, A. M., Tadić, V., Draganić, B., Ostafe, R., Fischer, R.,& Prodanović, R. (2019). Supplementary data for the article: Kostić, A. Ž.; Gašić, U. M.; Pešić, M. B.; Stanojević, S. P.; Barać, M. B.; Mačukanović-Jocić, M. P.; Avramov, S. N.; Tešić, Ž. L. Phytochemical Analysis and Total Antioxidant Capacity of Rhizome, Above-Ground Vegetative Parts and Flower of Three Iris Species. Chemistry and Biodiversity 2019, 16 (3), 1–17. https://doi.org/10.1002/cbdv.201800565.
Biochemical Engineering Journal
Elsevier..
Blažić M, Balaž AM, Tadić V, Draganić B, Ostafe R, Fischer R, Prodanović R. Supplementary data for the article: Kostić, A. Ž.; Gašić, U. M.; Pešić, M. B.; Stanojević, S. P.; Barać, M. B.; Mačukanović-Jocić, M. P.; Avramov, S. N.; Tešić, Ž. L. Phytochemical Analysis and Total Antioxidant Capacity of Rhizome, Above-Ground Vegetative Parts and Flower of Three Iris Species. Chemistry and Biodiversity 2019, 16 (3), 1–17. https://doi.org/10.1002/cbdv.201800565. Biochemical Engineering Journal. 2019;
Blažić Marija, Balaž Ana Marija, Tadić Vojin, Draganić Bojana, Ostafe Raluca, Fischer Rainer, Prodanović Radivoje, "Supplementary data for the article: Kostić, A. Ž.; Gašić, U. M.; Pešić, M. B.; Stanojević, S. P.; Barać, M. B.; Mačukanović-Jocić, M. P.; Avramov, S. N.; Tešić, Ž. L. Phytochemical Analysis and Total Antioxidant Capacity of Rhizome, Above-Ground Vegetative Parts and Flower of Three Iris Species. Chemistry and Biodiversity 2019, 16 (3), 1–17. https://doi.org/10.1002/cbdv.201800565" Biochemical Engineering Journal (2019)

Directed evolution of cellobiose dehydrogenase on the surface of yeast cells using resazurin-based fluorescent assay

Blažić, Marija; Balaž, Ana Marija; Prodanović, Olivera; Popović, Nikolina; Ostafe, Raluca; Fischer, Rainer; Prodanović, Radivoje

(Applied sciences, 2019)

TY  - JOUR
AU  - Blažić, Marija
AU  - Balaž, Ana Marija
AU  - Prodanović, Olivera
AU  - Popović, Nikolina
AU  - Ostafe, Raluca
AU  - Fischer, Rainer
AU  - Prodanović, Radivoje
PY  - 2019
UR  - http://cherry.chem.bg.ac.rs/handle/123456789/2909
AB  - 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 (10mMlactose and 0.08mMcellobiose), 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  - Applied sciences
T2  - Applied Sciences (Switzerland)
T1  - Directed evolution of cellobiose dehydrogenase on the surface of yeast cells using resazurin-based fluorescent assay
VL  - 9
IS  - 7
SP  - 1
EP  - 15
DO  - 10.3390/app9071413
ER  - 
@article{
author = "Blažić, Marija and Balaž, Ana Marija and Prodanović, Olivera and Popović, Nikolina and Ostafe, Raluca and Fischer, Rainer and Prodanović, Radivoje",
year = "2019",
url = "http://cherry.chem.bg.ac.rs/handle/123456789/2909",
abstract = "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 (10mMlactose and 0.08mMcellobiose), 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 = "Applied sciences",
journal = "Applied Sciences (Switzerland)",
title = "Directed evolution of cellobiose dehydrogenase on the surface of yeast cells using resazurin-based fluorescent assay",
volume = "9",
number = "7",
pages = "1-15",
doi = "10.3390/app9071413"
}
Blažić, 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.
Applied Sciences (Switzerland)
Applied sciences., 9(7), 1-15.
https://doi.org/10.3390/app9071413
Blažić 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. Applied Sciences (Switzerland). 2019;9(7):1-15
Blažić 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" Applied Sciences (Switzerland), 9, no. 7 (2019):1-15,
https://doi.org/10.3390/app9071413 .
6
4
6

Supplementary data for the article: Blažić, M.; Balaž, A. M.; 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. Applied Sciences (Switzerland) 2019, 9 (7). https://doi.org/10.3390/app9071413

Blažić, Marija; Balaž, Ana Marija; Prodanović, Olivera; Popović, Nikolina; Ostafe, Raluca; Fischer, Rainer; Prodanović, Radivoje

(Applied sciences, 2019)

TY  - BOOK
AU  - Blažić, Marija
AU  - Balaž, Ana Marija
AU  - Prodanović, Olivera
AU  - Popović, Nikolina
AU  - Ostafe, Raluca
AU  - Fischer, Rainer
AU  - Prodanović, Radivoje
PY  - 2019
UR  - http://cherry.chem.bg.ac.rs/handle/123456789/2912
PB  - Applied sciences
T2  - Applied Sciences (Switzerland)
T1  - Supplementary data for the article: Blažić, M.; Balaž, A. M.; 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. Applied Sciences (Switzerland) 2019, 9 (7). https://doi.org/10.3390/app9071413
ER  - 
@book{
author = "Blažić, Marija and Balaž, Ana Marija and Prodanović, Olivera and Popović, Nikolina and Ostafe, Raluca and Fischer, Rainer and Prodanović, Radivoje",
year = "2019",
url = "http://cherry.chem.bg.ac.rs/handle/123456789/2912",
publisher = "Applied sciences",
journal = "Applied Sciences (Switzerland)",
title = "Supplementary data for the article: Blažić, M.; Balaž, A. M.; 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. Applied Sciences (Switzerland) 2019, 9 (7). https://doi.org/10.3390/app9071413"
}
Blažić, M., Balaž, A. M., Prodanović, O., Popović, N., Ostafe, R., Fischer, R.,& Prodanović, R. (2019). Supplementary data for the article: Blažić, M.; Balaž, A. M.; 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. Applied Sciences (Switzerland) 2019, 9 (7). https://doi.org/10.3390/app9071413.
Applied Sciences (Switzerland)
Applied sciences..
Blažić M, Balaž AM, Prodanović O, Popović N, Ostafe R, Fischer R, Prodanović R. Supplementary data for the article: Blažić, M.; Balaž, A. M.; 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. Applied Sciences (Switzerland) 2019, 9 (7). https://doi.org/10.3390/app9071413. Applied Sciences (Switzerland). 2019;
Blažić Marija, Balaž Ana Marija, Prodanović Olivera, Popović Nikolina, Ostafe Raluca, Fischer Rainer, Prodanović Radivoje, "Supplementary data for the article: Blažić, M.; Balaž, A. M.; 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. Applied Sciences (Switzerland) 2019, 9 (7). https://doi.org/10.3390/app9071413" Applied Sciences (Switzerland) (2019)

Directed evolution of cellobiose dehydrogenase from Phanerochaete chrysosporium in yeast Saccharomyces cerevisiae for increased activity

Blažić, Marija; Prodanović, Radivoje

(Wiley, Hoboken, 2017)

TY  - CONF
AU  - Blažić, Marija
AU  - Prodanović, Radivoje
PY  - 2017
UR  - http://cherry.chem.bg.ac.rs/handle/123456789/2406
PB  - Wiley, Hoboken
C3  - FEBS Journal / Federation of European of Biochemical Societies
T1  - Directed evolution of cellobiose dehydrogenase from Phanerochaete chrysosporium in yeast Saccharomyces cerevisiae for increased activity
VL  - 284
SP  - 104
EP  - 104
ER  - 
@conference{
author = "Blažić, Marija and Prodanović, Radivoje",
year = "2017",
url = "http://cherry.chem.bg.ac.rs/handle/123456789/2406",
publisher = "Wiley, Hoboken",
journal = "FEBS Journal / Federation of European of Biochemical Societies",
title = "Directed evolution of cellobiose dehydrogenase from Phanerochaete chrysosporium in yeast Saccharomyces cerevisiae for increased activity",
volume = "284",
pages = "104-104"
}
Blažić, M.,& Prodanović, R. (2017). Directed evolution of cellobiose dehydrogenase from Phanerochaete chrysosporium in yeast Saccharomyces cerevisiae for increased activity.
FEBS Journal / Federation of European of Biochemical Societies
Wiley, Hoboken., 284, 104-104.
Blažić M, Prodanović R. Directed evolution of cellobiose dehydrogenase from Phanerochaete chrysosporium in yeast Saccharomyces cerevisiae for increased activity. FEBS Journal / Federation of European of Biochemical Societies. 2017;284:104-104
Blažić Marija, Prodanović Radivoje, "Directed evolution of cellobiose dehydrogenase from Phanerochaete chrysosporium in yeast Saccharomyces cerevisiae for increased activity" FEBS Journal / Federation of European of Biochemical Societies, 284 (2017):104-104

Supplementary data for article: Prodanović, O.; Spasojević, D.; Prokopijević, M.; Radotić, K.; Markovic, N.; Blažić, M.; Prodanović, R. Tyramine Modified Alginates via Periodate Oxidation for Peroxidase Induced Hydrogel Formation and Immobilization. Reactive and Functional Polymers 2015, 93, 77–83. https://doi.org/10.1016/j.reactfunctpolym.2015.06.004

Prodanović, Olivera; Spasojević, Dragica; Prokopijević, Miloš; Radotić, Ksenija; Marković, Nevena; Blažić, Marija; Prodanović, Radivoje

(Elsevier Science Bv, Amsterdam, 2015)

TY  - BOOK
AU  - Prodanović, Olivera
AU  - Spasojević, Dragica
AU  - Prokopijević, Miloš
AU  - Radotić, Ksenija
AU  - Marković, Nevena
AU  - Blažić, Marija
AU  - Prodanović, Radivoje
PY  - 2015
UR  - http://cherry.chem.bg.ac.rs/handle/123456789/3457
PB  - Elsevier Science Bv, Amsterdam
T2  - Reactive and Functional Polymers
T1  - Supplementary data for article: Prodanović, O.; Spasojević, D.; Prokopijević, M.; Radotić, K.; Markovic, N.; Blažić, M.; Prodanović, R. Tyramine Modified Alginates via Periodate Oxidation for Peroxidase Induced Hydrogel Formation and Immobilization. Reactive and Functional Polymers 2015, 93, 77–83. https://doi.org/10.1016/j.reactfunctpolym.2015.06.004
ER  - 
@book{
author = "Prodanović, Olivera and Spasojević, Dragica and Prokopijević, Miloš and Radotić, Ksenija and Marković, Nevena and Blažić, Marija and Prodanović, Radivoje",
year = "2015",
url = "http://cherry.chem.bg.ac.rs/handle/123456789/3457",
publisher = "Elsevier Science Bv, Amsterdam",
journal = "Reactive and Functional Polymers",
title = "Supplementary data for article: Prodanović, O.; Spasojević, D.; Prokopijević, M.; Radotić, K.; Markovic, N.; Blažić, M.; Prodanović, R. Tyramine Modified Alginates via Periodate Oxidation for Peroxidase Induced Hydrogel Formation and Immobilization. Reactive and Functional Polymers 2015, 93, 77–83. https://doi.org/10.1016/j.reactfunctpolym.2015.06.004"
}
Prodanović, O., Spasojević, D., Prokopijević, M., Radotić, K., Marković, N., Blažić, M.,& Prodanović, R. (2015). Supplementary data for article: Prodanović, O.; Spasojević, D.; Prokopijević, M.; Radotić, K.; Markovic, N.; Blažić, M.; Prodanović, R. Tyramine Modified Alginates via Periodate Oxidation for Peroxidase Induced Hydrogel Formation and Immobilization. Reactive and Functional Polymers 2015, 93, 77–83. https://doi.org/10.1016/j.reactfunctpolym.2015.06.004.
Reactive and Functional Polymers
Elsevier Science Bv, Amsterdam..
Prodanović O, Spasojević D, Prokopijević M, Radotić K, Marković N, Blažić M, Prodanović R. Supplementary data for article: Prodanović, O.; Spasojević, D.; Prokopijević, M.; Radotić, K.; Markovic, N.; Blažić, M.; Prodanović, R. Tyramine Modified Alginates via Periodate Oxidation for Peroxidase Induced Hydrogel Formation and Immobilization. Reactive and Functional Polymers 2015, 93, 77–83. https://doi.org/10.1016/j.reactfunctpolym.2015.06.004. Reactive and Functional Polymers. 2015;
Prodanović Olivera, Spasojević Dragica, Prokopijević Miloš, Radotić Ksenija, Marković Nevena, Blažić Marija, Prodanović Radivoje, "Supplementary data for article: Prodanović, O.; Spasojević, D.; Prokopijević, M.; Radotić, K.; Markovic, N.; Blažić, M.; Prodanović, R. Tyramine Modified Alginates via Periodate Oxidation for Peroxidase Induced Hydrogel Formation and Immobilization. Reactive and Functional Polymers 2015, 93, 77–83. https://doi.org/10.1016/j.reactfunctpolym.2015.06.004" Reactive and Functional Polymers (2015)

Tyramine modified alginates via periodate oxidation for peroxidase induced hydrogel formation and immobilization

Prodanović, Olivera; Spasojević, Dragica; Prokopijević, Miloš; Radotić, Ksenija; Marković, Nevena; Blažić, Marija; Prodanović, Radivoje

(Elsevier Science Bv, Amsterdam, 2015)

TY  - JOUR
AU  - Prodanović, Olivera
AU  - Spasojević, Dragica
AU  - Prokopijević, Miloš
AU  - Radotić, Ksenija
AU  - Marković, Nevena
AU  - Blažić, Marija
AU  - Prodanović, Radivoje
PY  - 2015
UR  - http://cherry.chem.bg.ac.rs/handle/123456789/1966
AB  - Phenol and amino groups were introduced into alginate to different degrees via oxidation with 2.5, 5, 10, 15 and 20 mol% of periodate and reductive amination by tyramine. Modification of alginate with tyramine was confirmed by FTIR spectroscopy and UV-VIS spectroscopy, while concentration of phenol and ionizable groups was determined using absorbance at 275 nm and acid-base titration. All tyramine-alginates were able to form hydrogels after cross-linking with horse radish peroxidase (HRP) and hydrogen peroxide. Tyramine-alginates oxidized with up to 10 mol% of periodate were also capable of forming hydrogels with calcium ions. Tyramine-alginates were tested for HRP immobilization within micro-beads obtained by peroxidase catalyzed droplet polymerization using internal delivery of hydrogen peroxide via glucose oxidase and glucose. Highest activity of immobilized peroxidase was obtained with 20% (w/v) tyramine-alginate obtained via 20 mol% periodate oxidation. Immobilized enzyme was not leaking from the micro-beads and was further kinetically characterized for pyrogallol oxidation. Km for pyrogallol was increased after immobilization from 1.93 mM for soluble HRP to 734 mM for immobilized HRP. The optimum pH was also increased from pH 7.0 to 8.0. Temperature and organic solvent stability improved significantly after immobilization, so that half-life at 70 degrees C increased around four times, while half-life in 80% (v/v) dioxane increased 22 times. After repeated use of 6 times in batch reactor for pyrogallol oxidation immobilized HRP retained 45% of original activity.
PB  - Elsevier Science Bv, Amsterdam
T2  - Reactive and Functional Polymers
T1  - Tyramine modified alginates via periodate oxidation for peroxidase induced hydrogel formation and immobilization
VL  - 93
SP  - 77
EP  - 83
DO  - 10.1016/j.reactfunctpolym.2015.06.004
ER  - 
@article{
author = "Prodanović, Olivera and Spasojević, Dragica and Prokopijević, Miloš and Radotić, Ksenija and Marković, Nevena and Blažić, Marija and Prodanović, Radivoje",
year = "2015",
url = "http://cherry.chem.bg.ac.rs/handle/123456789/1966",
abstract = "Phenol and amino groups were introduced into alginate to different degrees via oxidation with 2.5, 5, 10, 15 and 20 mol% of periodate and reductive amination by tyramine. Modification of alginate with tyramine was confirmed by FTIR spectroscopy and UV-VIS spectroscopy, while concentration of phenol and ionizable groups was determined using absorbance at 275 nm and acid-base titration. All tyramine-alginates were able to form hydrogels after cross-linking with horse radish peroxidase (HRP) and hydrogen peroxide. Tyramine-alginates oxidized with up to 10 mol% of periodate were also capable of forming hydrogels with calcium ions. Tyramine-alginates were tested for HRP immobilization within micro-beads obtained by peroxidase catalyzed droplet polymerization using internal delivery of hydrogen peroxide via glucose oxidase and glucose. Highest activity of immobilized peroxidase was obtained with 20% (w/v) tyramine-alginate obtained via 20 mol% periodate oxidation. Immobilized enzyme was not leaking from the micro-beads and was further kinetically characterized for pyrogallol oxidation. Km for pyrogallol was increased after immobilization from 1.93 mM for soluble HRP to 734 mM for immobilized HRP. The optimum pH was also increased from pH 7.0 to 8.0. Temperature and organic solvent stability improved significantly after immobilization, so that half-life at 70 degrees C increased around four times, while half-life in 80% (v/v) dioxane increased 22 times. After repeated use of 6 times in batch reactor for pyrogallol oxidation immobilized HRP retained 45% of original activity.",
publisher = "Elsevier Science Bv, Amsterdam",
journal = "Reactive and Functional Polymers",
title = "Tyramine modified alginates via periodate oxidation for peroxidase induced hydrogel formation and immobilization",
volume = "93",
pages = "77-83",
doi = "10.1016/j.reactfunctpolym.2015.06.004"
}
Prodanović, O., Spasojević, D., Prokopijević, M., Radotić, K., Marković, N., Blažić, M.,& Prodanović, R. (2015). Tyramine modified alginates via periodate oxidation for peroxidase induced hydrogel formation and immobilization.
Reactive and Functional Polymers
Elsevier Science Bv, Amsterdam., 93, 77-83.
https://doi.org/10.1016/j.reactfunctpolym.2015.06.004
Prodanović O, Spasojević D, Prokopijević M, Radotić K, Marković N, Blažić M, Prodanović R. Tyramine modified alginates via periodate oxidation for peroxidase induced hydrogel formation and immobilization. Reactive and Functional Polymers. 2015;93:77-83
Prodanović Olivera, Spasojević Dragica, Prokopijević Miloš, Radotić Ksenija, Marković Nevena, Blažić Marija, Prodanović Radivoje, "Tyramine modified alginates via periodate oxidation for peroxidase induced hydrogel formation and immobilization" Reactive and Functional Polymers, 93 (2015):77-83,
https://doi.org/10.1016/j.reactfunctpolym.2015.06.004 .
19
13
17

Expression of cellobiose dehydrogenase from Phanerochaete chrysosporium in yeast Saccharomyces cerevisiae for directed evolution

Blažić, Marija; Prodanović, Radivoje

(Wiley-Blackwell, Hoboken, 2015)

TY  - CONF
AU  - Blažić, Marija
AU  - Prodanović, Radivoje
PY  - 2015
UR  - http://cherry.chem.bg.ac.rs/handle/123456789/1975
PB  - Wiley-Blackwell, Hoboken
C3  - FEBS Journal / Federation of European of Biochemical Societies
T1  - Expression of cellobiose dehydrogenase from Phanerochaete chrysosporium in yeast Saccharomyces cerevisiae for directed evolution
VL  - 282
SP  - 119
EP  - 119
ER  - 
@conference{
author = "Blažić, Marija and Prodanović, Radivoje",
year = "2015",
url = "http://cherry.chem.bg.ac.rs/handle/123456789/1975",
publisher = "Wiley-Blackwell, Hoboken",
journal = "FEBS Journal / Federation of European of Biochemical Societies",
title = "Expression of cellobiose dehydrogenase from Phanerochaete chrysosporium in yeast Saccharomyces cerevisiae for directed evolution",
volume = "282",
pages = "119-119"
}
Blažić, M.,& Prodanović, R. (2015). Expression of cellobiose dehydrogenase from Phanerochaete chrysosporium in yeast Saccharomyces cerevisiae for directed evolution.
FEBS Journal / Federation of European of Biochemical Societies
Wiley-Blackwell, Hoboken., 282, 119-119.
Blažić M, Prodanović R. Expression of cellobiose dehydrogenase from Phanerochaete chrysosporium in yeast Saccharomyces cerevisiae for directed evolution. FEBS Journal / Federation of European of Biochemical Societies. 2015;282:119-119
Blažić Marija, Prodanović Radivoje, "Expression of cellobiose dehydrogenase from Phanerochaete chrysosporium in yeast Saccharomyces cerevisiae for directed evolution" FEBS Journal / Federation of European of Biochemical Societies, 282 (2015):119-119

Cloning, Heterologous Expression, Purification and Characterization of M12 Mutant of Aspergillus niger Glucose Oxidase in Yeast Pichia pastoris KM71H

Kovačević, Gordana; Blažić, Marija; Draganić, Bojana; Ostafe, Raluca; Gavrović-Jankulović, Marija; Fischer, Rainer; Prodanović, Radivoje

(Humana Press Inc, Totowa, 2014)

TY  - JOUR
AU  - Kovačević, Gordana
AU  - Blažić, Marija
AU  - Draganić, Bojana
AU  - Ostafe, Raluca
AU  - Gavrović-Jankulović, Marija
AU  - Fischer, Rainer
AU  - Prodanović, Radivoje
PY  - 2014
UR  - http://cherry.chem.bg.ac.rs/handle/123456789/1517
AB  - Aspergillus niger glucose oxidase (GOx) genes for wild-type (GenBank accession no. X16061, swiss-Prot; P13006) and M12 mutant (N2Y, K13E, T30 V, I94 V, K152R) were cloned into pPICZ alpha A vector for expression in Pichia pastoris KM71H strain. The highest expression level of 17.5 U/mL of fermentation media was obtained in 0.5 % (v/v) methanol after 9 days of fermentation. The recombinant GOx was purified by cross-flow ultrafiltration using membranes of 30 kDa molecular cutoff and DEAE ion-exchange chromatography at pH 6.0. Purified wt GOx had k (cat) of 189.4 s(-1) and K (m) of 28.26 mM while M12 GOx had k (cat) of 352.0 s(-1) and K (m) of 13.33 mM for glucose at pH 5.5. Specificity constants k (cat)/K (m) of wt (6.70 mM(-1) s(-1)) and M12 GOx (26.7 mM(-1) s(-1)) expressed in P. pastoris KM71H were around three times higher than for the same enzymes previously expressed in Saccharomyces cerevisiae InvSc1 strain. The pH optimum and sugar specificity of M12 mutant of GOx remained similar to the wild-type form of the enzyme, while thermostability was slightly decreased. M12 GOx expressed in P. pastoris showed three times higher activity compared to the wt GOx toward redox mediators like N,N-dimethyl-nitroso-aniline used for glucose strips manufacturing. M12 mutant of GOx produced in P. pastoris KM71H could be useful for manufacturing of glucose biosensors and biofuel cells.
PB  - Humana Press Inc, Totowa
T2  - Molecular Biotechnology
T1  - Cloning, Heterologous Expression, Purification and Characterization of M12 Mutant of Aspergillus niger Glucose Oxidase in Yeast Pichia pastoris KM71H
VL  - 56
IS  - 4
SP  - 305
EP  - 311
DO  - 10.1007/s12033-013-9709-x
ER  - 
@article{
author = "Kovačević, Gordana and Blažić, Marija and Draganić, Bojana and Ostafe, Raluca and Gavrović-Jankulović, Marija and Fischer, Rainer and Prodanović, Radivoje",
year = "2014",
url = "http://cherry.chem.bg.ac.rs/handle/123456789/1517",
abstract = "Aspergillus niger glucose oxidase (GOx) genes for wild-type (GenBank accession no. X16061, swiss-Prot; P13006) and M12 mutant (N2Y, K13E, T30 V, I94 V, K152R) were cloned into pPICZ alpha A vector for expression in Pichia pastoris KM71H strain. The highest expression level of 17.5 U/mL of fermentation media was obtained in 0.5 % (v/v) methanol after 9 days of fermentation. The recombinant GOx was purified by cross-flow ultrafiltration using membranes of 30 kDa molecular cutoff and DEAE ion-exchange chromatography at pH 6.0. Purified wt GOx had k (cat) of 189.4 s(-1) and K (m) of 28.26 mM while M12 GOx had k (cat) of 352.0 s(-1) and K (m) of 13.33 mM for glucose at pH 5.5. Specificity constants k (cat)/K (m) of wt (6.70 mM(-1) s(-1)) and M12 GOx (26.7 mM(-1) s(-1)) expressed in P. pastoris KM71H were around three times higher than for the same enzymes previously expressed in Saccharomyces cerevisiae InvSc1 strain. The pH optimum and sugar specificity of M12 mutant of GOx remained similar to the wild-type form of the enzyme, while thermostability was slightly decreased. M12 GOx expressed in P. pastoris showed three times higher activity compared to the wt GOx toward redox mediators like N,N-dimethyl-nitroso-aniline used for glucose strips manufacturing. M12 mutant of GOx produced in P. pastoris KM71H could be useful for manufacturing of glucose biosensors and biofuel cells.",
publisher = "Humana Press Inc, Totowa",
journal = "Molecular Biotechnology",
title = "Cloning, Heterologous Expression, Purification and Characterization of M12 Mutant of Aspergillus niger Glucose Oxidase in Yeast Pichia pastoris KM71H",
volume = "56",
number = "4",
pages = "305-311",
doi = "10.1007/s12033-013-9709-x"
}
Kovačević, G., Blažić, M., Draganić, B., Ostafe, R., Gavrović-Jankulović, M., Fischer, R.,& Prodanović, R. (2014). Cloning, Heterologous Expression, Purification and Characterization of M12 Mutant of Aspergillus niger Glucose Oxidase in Yeast Pichia pastoris KM71H.
Molecular Biotechnology
Humana Press Inc, Totowa., 56(4), 305-311.
https://doi.org/10.1007/s12033-013-9709-x
Kovačević G, Blažić M, Draganić B, Ostafe R, Gavrović-Jankulović M, Fischer R, Prodanović R. Cloning, Heterologous Expression, Purification and Characterization of M12 Mutant of Aspergillus niger Glucose Oxidase in Yeast Pichia pastoris KM71H. Molecular Biotechnology. 2014;56(4):305-311
Kovačević Gordana, Blažić Marija, Draganić Bojana, Ostafe Raluca, Gavrović-Jankulović Marija, Fischer Rainer, Prodanović Radivoje, "Cloning, Heterologous Expression, Purification and Characterization of M12 Mutant of Aspergillus niger Glucose Oxidase in Yeast Pichia pastoris KM71H" Molecular Biotechnology, 56, no. 4 (2014):305-311,
https://doi.org/10.1007/s12033-013-9709-x .
18
17
19

Supplementary data for article: Blažić, M.; Kovačević, G.; Prodanović, O.; Ostafe, R.; Gavrović-Jankulović, M.; Fischer, R.; Prodanović, R. Yeast Surface Display for the Expression, Purification and Characterization of Wild-Type and B11 Mutant Glucose Oxidases. Protein Expression and Purification 2013, 89 (2), 175–180. https://doi.org/10.1016/j.pep.2013.03.014

Blažić, Marija; Kovačević, Gordana; Prodanović, Olivera; Ostafe, Raluca; Gavrović-Jankulović, Marija; Fischer, Rainer; Prodanović, Radivoje

(Academic Press Inc Elsevier Science, San Diego, 2013)

TY  - BOOK
AU  - Blažić, Marija
AU  - Kovačević, Gordana
AU  - Prodanović, Olivera
AU  - Ostafe, Raluca
AU  - Gavrović-Jankulović, Marija
AU  - Fischer, Rainer
AU  - Prodanović, Radivoje
PY  - 2013
UR  - http://cherry.chem.bg.ac.rs/handle/123456789/3568
PB  - Academic Press Inc Elsevier Science, San Diego
T2  - Protein Expression and Purification
T1  - Supplementary data for article: Blažić, M.; Kovačević, G.; Prodanović, O.; Ostafe, R.; Gavrović-Jankulović, M.; Fischer, R.; Prodanović, R. Yeast Surface Display for the Expression, Purification and Characterization of Wild-Type and B11 Mutant Glucose Oxidases. Protein Expression and Purification 2013, 89 (2), 175–180. https://doi.org/10.1016/j.pep.2013.03.014
ER  - 
@book{
author = "Blažić, Marija and Kovačević, Gordana and Prodanović, Olivera and Ostafe, Raluca and Gavrović-Jankulović, Marija and Fischer, Rainer and Prodanović, Radivoje",
year = "2013",
url = "http://cherry.chem.bg.ac.rs/handle/123456789/3568",
publisher = "Academic Press Inc Elsevier Science, San Diego",
journal = "Protein Expression and Purification",
title = "Supplementary data for article: Blažić, M.; Kovačević, G.; Prodanović, O.; Ostafe, R.; Gavrović-Jankulović, M.; Fischer, R.; Prodanović, R. Yeast Surface Display for the Expression, Purification and Characterization of Wild-Type and B11 Mutant Glucose Oxidases. Protein Expression and Purification 2013, 89 (2), 175–180. https://doi.org/10.1016/j.pep.2013.03.014"
}
Blažić, M., Kovačević, G., Prodanović, O., Ostafe, R., Gavrović-Jankulović, M., Fischer, R.,& Prodanović, R. (2013). Supplementary data for article: Blažić, M.; Kovačević, G.; Prodanović, O.; Ostafe, R.; Gavrović-Jankulović, M.; Fischer, R.; Prodanović, R. Yeast Surface Display for the Expression, Purification and Characterization of Wild-Type and B11 Mutant Glucose Oxidases. Protein Expression and Purification 2013, 89 (2), 175–180. https://doi.org/10.1016/j.pep.2013.03.014.
Protein Expression and Purification
Academic Press Inc Elsevier Science, San Diego..
Blažić M, Kovačević G, Prodanović O, Ostafe R, Gavrović-Jankulović M, Fischer R, Prodanović R. Supplementary data for article: Blažić, M.; Kovačević, G.; Prodanović, O.; Ostafe, R.; Gavrović-Jankulović, M.; Fischer, R.; Prodanović, R. Yeast Surface Display for the Expression, Purification and Characterization of Wild-Type and B11 Mutant Glucose Oxidases. Protein Expression and Purification 2013, 89 (2), 175–180. https://doi.org/10.1016/j.pep.2013.03.014. Protein Expression and Purification. 2013;
Blažić Marija, Kovačević Gordana, Prodanović Olivera, Ostafe Raluca, Gavrović-Jankulović Marija, Fischer Rainer, Prodanović Radivoje, "Supplementary data for article: Blažić, M.; Kovačević, G.; Prodanović, O.; Ostafe, R.; Gavrović-Jankulović, M.; Fischer, R.; Prodanović, R. Yeast Surface Display for the Expression, Purification and Characterization of Wild-Type and B11 Mutant Glucose Oxidases. Protein Expression and Purification 2013, 89 (2), 175–180. https://doi.org/10.1016/j.pep.2013.03.014" Protein Expression and Purification (2013)

Yeast surface display for the expression, purification and characterization of wild-type and B11 mutant glucose oxidases

Blažić, Marija; Kovačević, Gordana; Prodanović, Olivera; Ostafe, Raluca; Gavrović-Jankulović, Marija; Fischer, Rainer; Prodanović, Radivoje

(Academic Press Inc Elsevier Science, San Diego, 2013)

TY  - JOUR
AU  - Blažić, Marija
AU  - Kovačević, Gordana
AU  - Prodanović, Olivera
AU  - Ostafe, Raluca
AU  - Gavrović-Jankulović, Marija
AU  - Fischer, Rainer
AU  - Prodanović, Radivoje
PY  - 2013
UR  - http://cherry.chem.bg.ac.rs/handle/123456789/1357
AB  - Glucose oxidase (GOx) catalyzes the oxidation of glucose to form gluconic acid and hydrogen peroxide, a reaction with important applications in food preservation, the manufacture of cosmetics and pharmaceuticals, and the development of glucose monitoring devices and biofuel cells. We expressed Aspergillus niger wild type GOx and the B11 mutant, which has twice the activity of the wild type enzyme at pH 5.5, as C-terminal fusions with the Saccharomyces cerevisiae Aga2 protein, allowing the fusion proteins to be displayed on the surface of yeast EBY100 cells. After expression, we extracted the proteins from the yeast cell wall and purified them by ion-exchange chromatography and ultrafiltration. This produced a broad 100-140 kDa band by denaturing SDS-PAGE and a high-molecular-weight band by native PAGE corresponding to the activity band revealed by zymography. The wild type and B11 fusion proteins had k(cat) values of 33.3 and 61.3 s(-1) and K-m values for glucose of 33.4 and 27.9 mM, respectively. The pH optimum for both enzymes was 5.0. The kinetic properties of the fusion proteins displayed the same ratio as their native counterparts, confirming that yeast surface display is suitable for the high-throughput directed evolution of GOx using flow cytometry for selection. Aga2-GOx fusion proteins in the yeast cell wall could also be used as immobilized catalysts for the production of gluconic acid.
PB  - Academic Press Inc Elsevier Science, San Diego
T2  - Protein Expression and Purification
T1  - Yeast surface display for the expression, purification and characterization of wild-type and B11 mutant glucose oxidases
VL  - 89
IS  - 2
SP  - 175
EP  - 180
DO  - 10.1016/j.pep.2013.03.014
ER  - 
@article{
author = "Blažić, Marija and Kovačević, Gordana and Prodanović, Olivera and Ostafe, Raluca and Gavrović-Jankulović, Marija and Fischer, Rainer and Prodanović, Radivoje",
year = "2013",
url = "http://cherry.chem.bg.ac.rs/handle/123456789/1357",
abstract = "Glucose oxidase (GOx) catalyzes the oxidation of glucose to form gluconic acid and hydrogen peroxide, a reaction with important applications in food preservation, the manufacture of cosmetics and pharmaceuticals, and the development of glucose monitoring devices and biofuel cells. We expressed Aspergillus niger wild type GOx and the B11 mutant, which has twice the activity of the wild type enzyme at pH 5.5, as C-terminal fusions with the Saccharomyces cerevisiae Aga2 protein, allowing the fusion proteins to be displayed on the surface of yeast EBY100 cells. After expression, we extracted the proteins from the yeast cell wall and purified them by ion-exchange chromatography and ultrafiltration. This produced a broad 100-140 kDa band by denaturing SDS-PAGE and a high-molecular-weight band by native PAGE corresponding to the activity band revealed by zymography. The wild type and B11 fusion proteins had k(cat) values of 33.3 and 61.3 s(-1) and K-m values for glucose of 33.4 and 27.9 mM, respectively. The pH optimum for both enzymes was 5.0. The kinetic properties of the fusion proteins displayed the same ratio as their native counterparts, confirming that yeast surface display is suitable for the high-throughput directed evolution of GOx using flow cytometry for selection. Aga2-GOx fusion proteins in the yeast cell wall could also be used as immobilized catalysts for the production of gluconic acid.",
publisher = "Academic Press Inc Elsevier Science, San Diego",
journal = "Protein Expression and Purification",
title = "Yeast surface display for the expression, purification and characterization of wild-type and B11 mutant glucose oxidases",
volume = "89",
number = "2",
pages = "175-180",
doi = "10.1016/j.pep.2013.03.014"
}
Blažić, M., Kovačević, G., Prodanović, O., Ostafe, R., Gavrović-Jankulović, M., Fischer, R.,& Prodanović, R. (2013). Yeast surface display for the expression, purification and characterization of wild-type and B11 mutant glucose oxidases.
Protein Expression and Purification
Academic Press Inc Elsevier Science, San Diego., 89(2), 175-180.
https://doi.org/10.1016/j.pep.2013.03.014
Blažić M, Kovačević G, Prodanović O, Ostafe R, Gavrović-Jankulović M, Fischer R, Prodanović R. Yeast surface display for the expression, purification and characterization of wild-type and B11 mutant glucose oxidases. Protein Expression and Purification. 2013;89(2):175-180
Blažić Marija, Kovačević Gordana, Prodanović Olivera, Ostafe Raluca, Gavrović-Jankulović Marija, Fischer Rainer, Prodanović Radivoje, "Yeast surface display for the expression, purification and characterization of wild-type and B11 mutant glucose oxidases" Protein Expression and Purification, 89, no. 2 (2013):175-180,
https://doi.org/10.1016/j.pep.2013.03.014 .
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Nanobiocatalysts for biofuel cells and biosensor systems

Prodanović, Radivoje; Gavrović-Jankulović, Marija; Kovačević, Gordana; Blažić, Marija; Prodanović, Olivera; Ostafe, Raluca

(2011)

TY  - JOUR
AU  - Prodanović, Radivoje
AU  - Gavrović-Jankulović, Marija
AU  - Kovačević, Gordana
AU  - Blažić, Marija
AU  - Prodanović, Olivera
AU  - Ostafe, Raluca
PY  - 2011
UR  - http://cherry.chem.bg.ac.rs/handle/123456789/135
AB  - This overview summarizes the application of enzymes in the manufacture and design of biofuel cells and biosensors. The emphasis will be put on the protein engineering techniques used for improving the properties of enzymes such as nanobiocatalysts, e.g. immobilization orientation, stability, activity and efficiency of electron transfer between immobilized enzymes and electrodes. Some possible applications in the military and some future designs of these electric devices will be discussed as well.
AB  - U ovom preglednom članku je sumirana primena enzima u proizvodnji i dizajnu biogorivnih ćelija i biosenzora. Naglasak u pregledu literature je stavljen na tehnike proteinskog inžinjeringa, koje se koriste za poboljšanje osobina enzima u nanobiokatalizatorima kao što su orijentacija kod imobilizacije, stabilnost, aktivnost i efikasnost transfera elektrona između imobilizovanog enzima i elektrode. Na kraju pregleda je dato nekoliko primera moguće primene u vojsci. Nanobiokatalizatori su biokatalizatori u obliku enzima ili ćelija imobilizovani na nanomaterijalima. Koriste se kao sastavni elementi gorivnih ćelija u vidu imobilizovanih oksidoreduktaza na elektrodama. Na anodi se uz pomoć enzima oksiduju hemijska jedinjenja i elektroni predaju elektrodi, dok se na katodi elektroni uz pomoć druge oksidoreduktaze prebacuju sa elektrode na vodu ili kiseonik. Enzimi koji se koriste na anodi su glukoza oksidaza, formaldehid dehidrogenaza, alkohol dehidrogenaza i druge oksidaze šećera. Na katodi se uglavnom koriste lakaze, bilirubin oksidaza, peroksidaze i citohrom c oksidaza. Zahvaljujući razvoju nanotehnologije razvijaju se i minijaturne biogorivne ćelije koje proizvode električnu energiju za implantirane medicinske uređaje (insulinske pumpe, pejsmejkere, biosenzore) koristeći glukozu i kiseonik iz ljudske krvi. Biosenzori predstavljaju uređaje koji se sastoje iz biološke komponente, transducera i električne komponente. Oni pretvaraju koncen traciju hemijske supstance u električni signal i koriste se za analitiku. Kao biološka komponenta se mogu koristiti enzimi, monoklonska antitela, nukleinske kiseline i lipidi. Enzimska logička kola predstavljaju kombinaciju različitih biosenzora (enzimskih reakcija) koji mere nekoliko ulaznih parametara i na osnovu njih daju odgovarajući izlazni signal. Koristeći znanja kompjuterske tehnologije enzimskim logičkim kolima mogu se simulirati AND, OR, XOR, NOR, NAND, INHIB i XNOR logička kola. Za poboljšanje osobina biokatalizatora u cilju efikasnije primene u bioelektrokatalizi koriste se tehnike proteinskog inžinjeringa kao što su racionalni dizajn i dirigovana evolucija. Dirigovana evolucija koristi iterativne korake mutiranja i selekcije, kako bi biokatalizator evoluirao u pravcu koji nam je potreban. Najsporiji stupanj u ovoj tehnologiji predstavlja 'skrining', te se u novije vreme pomoću protočne citometrije i mikrofluidike pokušavaju razviti nove metode visoko propusnog skrininga. U literaturi opisani primeri dirigovane evolucije glukoza oksidaze, glukoza dehidrogenaze, formaldehid dehidrogenaze, laktat dehidrogenaze, peroksidaze i lakaze. Kombinacijom enzimskih logičkih kola i mikrofluidne tehnologije se pokušavaju napraviti laboratorije na čipu koje bi omogućile kontinuirano praćenje zdravstvenog stanja vojnika na bojnom polju i u slučaju šoka (ranjavanja) primenu odgovarajuće terapije u toku prvih 30 minuta od povrede. To bi obezbedilo veći stepen preživljavanja vojnika u ratu. Takođe upotrebom enzimskih logičkih kola i antitela moguće je postići uskladištenje i šifrovanje informacija, kao i zaštitu lozinkom, odgovarajućih elektronskih uređaja kao što su biogorivne ćelije Razvoj nanotehnologije, proteinskog inžinjeringa i molekularnog računarstva otvara vrata novim mogućnostima u proizvodnji i dizajnu biogorivnih ćelija i bisenzorskih sistema, kao i u skladištenju i zaštiti informacija.
T2  - Vojnotehnički glasnik
T1  - Nanobiocatalysts for biofuel cells and biosensor systems
T1  - Nanobiokatalizatori za biogorivne ćelije i biosenzorne sisteme
VL  - 59
IS  - 4
SP  - 79
EP  - 92
DO  - 10.5937/vojtehg1104079P
ER  - 
@article{
author = "Prodanović, Radivoje and Gavrović-Jankulović, Marija and Kovačević, Gordana and Blažić, Marija and Prodanović, Olivera and Ostafe, Raluca",
year = "2011",
url = "http://cherry.chem.bg.ac.rs/handle/123456789/135",
abstract = "This overview summarizes the application of enzymes in the manufacture and design of biofuel cells and biosensors. The emphasis will be put on the protein engineering techniques used for improving the properties of enzymes such as nanobiocatalysts, e.g. immobilization orientation, stability, activity and efficiency of electron transfer between immobilized enzymes and electrodes. Some possible applications in the military and some future designs of these electric devices will be discussed as well., U ovom preglednom članku je sumirana primena enzima u proizvodnji i dizajnu biogorivnih ćelija i biosenzora. Naglasak u pregledu literature je stavljen na tehnike proteinskog inžinjeringa, koje se koriste za poboljšanje osobina enzima u nanobiokatalizatorima kao što su orijentacija kod imobilizacije, stabilnost, aktivnost i efikasnost transfera elektrona između imobilizovanog enzima i elektrode. Na kraju pregleda je dato nekoliko primera moguće primene u vojsci. Nanobiokatalizatori su biokatalizatori u obliku enzima ili ćelija imobilizovani na nanomaterijalima. Koriste se kao sastavni elementi gorivnih ćelija u vidu imobilizovanih oksidoreduktaza na elektrodama. Na anodi se uz pomoć enzima oksiduju hemijska jedinjenja i elektroni predaju elektrodi, dok se na katodi elektroni uz pomoć druge oksidoreduktaze prebacuju sa elektrode na vodu ili kiseonik. Enzimi koji se koriste na anodi su glukoza oksidaza, formaldehid dehidrogenaza, alkohol dehidrogenaza i druge oksidaze šećera. Na katodi se uglavnom koriste lakaze, bilirubin oksidaza, peroksidaze i citohrom c oksidaza. Zahvaljujući razvoju nanotehnologije razvijaju se i minijaturne biogorivne ćelije koje proizvode električnu energiju za implantirane medicinske uređaje (insulinske pumpe, pejsmejkere, biosenzore) koristeći glukozu i kiseonik iz ljudske krvi. Biosenzori predstavljaju uređaje koji se sastoje iz biološke komponente, transducera i električne komponente. Oni pretvaraju koncen traciju hemijske supstance u električni signal i koriste se za analitiku. Kao biološka komponenta se mogu koristiti enzimi, monoklonska antitela, nukleinske kiseline i lipidi. Enzimska logička kola predstavljaju kombinaciju različitih biosenzora (enzimskih reakcija) koji mere nekoliko ulaznih parametara i na osnovu njih daju odgovarajući izlazni signal. Koristeći znanja kompjuterske tehnologije enzimskim logičkim kolima mogu se simulirati AND, OR, XOR, NOR, NAND, INHIB i XNOR logička kola. Za poboljšanje osobina biokatalizatora u cilju efikasnije primene u bioelektrokatalizi koriste se tehnike proteinskog inžinjeringa kao što su racionalni dizajn i dirigovana evolucija. Dirigovana evolucija koristi iterativne korake mutiranja i selekcije, kako bi biokatalizator evoluirao u pravcu koji nam je potreban. Najsporiji stupanj u ovoj tehnologiji predstavlja 'skrining', te se u novije vreme pomoću protočne citometrije i mikrofluidike pokušavaju razviti nove metode visoko propusnog skrininga. U literaturi opisani primeri dirigovane evolucije glukoza oksidaze, glukoza dehidrogenaze, formaldehid dehidrogenaze, laktat dehidrogenaze, peroksidaze i lakaze. Kombinacijom enzimskih logičkih kola i mikrofluidne tehnologije se pokušavaju napraviti laboratorije na čipu koje bi omogućile kontinuirano praćenje zdravstvenog stanja vojnika na bojnom polju i u slučaju šoka (ranjavanja) primenu odgovarajuće terapije u toku prvih 30 minuta od povrede. To bi obezbedilo veći stepen preživljavanja vojnika u ratu. Takođe upotrebom enzimskih logičkih kola i antitela moguće je postići uskladištenje i šifrovanje informacija, kao i zaštitu lozinkom, odgovarajućih elektronskih uređaja kao što su biogorivne ćelije Razvoj nanotehnologije, proteinskog inžinjeringa i molekularnog računarstva otvara vrata novim mogućnostima u proizvodnji i dizajnu biogorivnih ćelija i bisenzorskih sistema, kao i u skladištenju i zaštiti informacija.",
journal = "Vojnotehnički glasnik",
title = "Nanobiocatalysts for biofuel cells and biosensor systems, Nanobiokatalizatori za biogorivne ćelije i biosenzorne sisteme",
volume = "59",
number = "4",
pages = "79-92",
doi = "10.5937/vojtehg1104079P"
}
Prodanović, R., Gavrović-Jankulović, M., Kovačević, G., Blažić, M., Prodanović, O.,& Ostafe, R. (2011). Nanobiokatalizatori za biogorivne ćelije i biosenzorne sisteme.
Vojnotehnički glasnik, 59(4), 79-92.
https://doi.org/10.5937/vojtehg1104079P
Prodanović R, Gavrović-Jankulović M, Kovačević G, Blažić M, Prodanović O, Ostafe R. Nanobiokatalizatori za biogorivne ćelije i biosenzorne sisteme. Vojnotehnički glasnik. 2011;59(4):79-92
Prodanović Radivoje, Gavrović-Jankulović Marija, Kovačević Gordana, Blažić Marija, Prodanović Olivera, Ostafe Raluca, "Nanobiokatalizatori za biogorivne ćelije i biosenzorne sisteme" Vojnotehnički glasnik, 59, no. 4 (2011):79-92,
https://doi.org/10.5937/vojtehg1104079P .