Zelenović, Nevena D.

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  • Zelenović, Nevena D. (2)
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Author's Bibliography

Peroxidase-Sensitive Tyramine Carboxymethyl Xylan Hydrogels for Enzyme Encapsulation

Spasojević, Dragica; Prokopijević, Miloš; Prodanović, Olivera; Zelenović, Nevena D.; Polović, Natalija; Radotić, Ksenija; Prodanović, Radivoje

(The Polymer Society of Korea, 2019)

TY  - JOUR
AU  - Spasojević, Dragica
AU  - Prokopijević, Miloš
AU  - Prodanović, Olivera
AU  - Zelenović, Nevena D.
AU  - Polović, Natalija
AU  - Radotić, Ksenija
AU  - Prodanović, Radivoje
PY  - 2019
UR  - http://cherry.chem.bg.ac.rs/handle/123456789/3732
AB  - Derivatives of xylans were synthesized from corncob xylan by carboxymethylation, oxidization with different molar ratios of periodate (5, 10 15 and 20 mol%) and by reductive amination with tyramine. Modifications of tyramine carboxymethyl xylans (Tyr-CMX) were confirmed by FTIR, UV and NMR spectra. Concentration of ionizable groups increased from 1.5 mmol/g for carboxymethyl xylan (CMX) to 5.4 mmol/g for Tyr-CMX oxidized with 20 mol% of periodate. All Tyr-CMXs were able to form hydrogels the cross-linking reaction with horseradish peroxidase and peroxide. Tyr-CMXs were tested for amyloglucosidase (AG) encapsulation within hydrogel microbeads obtained in a reaction of emulsion polymerization with peroxidase. Average diameter of Tyr-CMX hydrogel microbeads was 52±25 µm and after encapsulation optimization with respect to the extent of CMX modification with tyramine, the concentration of Tyr-CMX, and the amount of added AG, microbeads with AG specific activity of 2 U/mL and 20% yield of immobilization were obtained. The optimum pH of the immobilized AG was not changed compared to the soluble one, while half-life at 60 °C was increased around 10 times. The Michaelis-Menten constant for the immobilized enzyme, 1.03 mM, was significantly lower than that for the soluble one, 1.54 mM. After 5 cycles of repetitive use in batch reactor, the immobilized AG retained 68% of initial activity.
PB  - The Polymer Society of Korea
T2  - Macromolecular Research
T1  - Peroxidase-Sensitive Tyramine Carboxymethyl Xylan Hydrogels for Enzyme Encapsulation
VL  - 27
IS  - 8
SP  - 764
EP  - 771
DO  - 10.1007/s13233-019-7111-7
ER  - 
@article{
author = "Spasojević, Dragica and Prokopijević, Miloš and Prodanović, Olivera and Zelenović, Nevena D. and Polović, Natalija and Radotić, Ksenija and Prodanović, Radivoje",
year = "2019",
url = "http://cherry.chem.bg.ac.rs/handle/123456789/3732",
abstract = "Derivatives of xylans were synthesized from corncob xylan by carboxymethylation, oxidization with different molar ratios of periodate (5, 10 15 and 20 mol%) and by reductive amination with tyramine. Modifications of tyramine carboxymethyl xylans (Tyr-CMX) were confirmed by FTIR, UV and NMR spectra. Concentration of ionizable groups increased from 1.5 mmol/g for carboxymethyl xylan (CMX) to 5.4 mmol/g for Tyr-CMX oxidized with 20 mol% of periodate. All Tyr-CMXs were able to form hydrogels the cross-linking reaction with horseradish peroxidase and peroxide. Tyr-CMXs were tested for amyloglucosidase (AG) encapsulation within hydrogel microbeads obtained in a reaction of emulsion polymerization with peroxidase. Average diameter of Tyr-CMX hydrogel microbeads was 52±25 µm and after encapsulation optimization with respect to the extent of CMX modification with tyramine, the concentration of Tyr-CMX, and the amount of added AG, microbeads with AG specific activity of 2 U/mL and 20% yield of immobilization were obtained. The optimum pH of the immobilized AG was not changed compared to the soluble one, while half-life at 60 °C was increased around 10 times. The Michaelis-Menten constant for the immobilized enzyme, 1.03 mM, was significantly lower than that for the soluble one, 1.54 mM. After 5 cycles of repetitive use in batch reactor, the immobilized AG retained 68% of initial activity.",
publisher = "The Polymer Society of Korea",
journal = "Macromolecular Research",
title = "Peroxidase-Sensitive Tyramine Carboxymethyl Xylan Hydrogels for Enzyme Encapsulation",
volume = "27",
number = "8",
pages = "764-771",
doi = "10.1007/s13233-019-7111-7"
}
Spasojević, D., Prokopijević, M., Prodanović, O., Zelenović, N. D., Polović, N., Radotić, K.,& Prodanović, R. (2019). Peroxidase-Sensitive Tyramine Carboxymethyl Xylan Hydrogels for Enzyme Encapsulation.
Macromolecular Research
The Polymer Society of Korea., 27(8), 764-771.
https://doi.org/10.1007/s13233-019-7111-7
Spasojević D, Prokopijević M, Prodanović O, Zelenović ND, Polović N, Radotić K, Prodanović R. Peroxidase-Sensitive Tyramine Carboxymethyl Xylan Hydrogels for Enzyme Encapsulation. Macromolecular Research. 2019;27(8):764-771
Spasojević Dragica, Prokopijević Miloš, Prodanović Olivera, Zelenović Nevena D., Polović Natalija, Radotić Ksenija, Prodanović Radivoje, "Peroxidase-Sensitive Tyramine Carboxymethyl Xylan Hydrogels for Enzyme Encapsulation" Macromolecular Research, 27, no. 8 (2019):764-771,
https://doi.org/10.1007/s13233-019-7111-7 .
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Cloning of the gene for a carbohydrate oxidase from Lactuca sativa in the yeasts Saccharomyces cerevisiae and Pichia pastoris

Tadić, Vojin; Balaž, Ana Marija; Petric, Marija P.; Milošević, Snežana; Zelenović, Nevena D.; Raspor, Martin Z.; Tadić, Jovan; Prodanović, Radivoje

(Assoc Chemical Engineers Serbia, Belgrade, 2015)

TY  - JOUR
AU  - Tadić, Vojin
AU  - Balaž, Ana Marija
AU  - Petric, Marija P.
AU  - Milošević, Snežana
AU  - Zelenović, Nevena D.
AU  - Raspor, Martin Z.
AU  - Tadić, Jovan
AU  - Prodanović, Radivoje
PY  - 2015
UR  - http://cherry.chem.bg.ac.rs/handle/123456789/2040
AB  - We have cloned the gene for carbohydrate oxidase (CHO) from Lactuca sativa in two species of yeasts (Saccharomyces cerevisiae and Pichia pastoris). The synthetic gene for the carbohydrate oxidase (1821 bp) from L. sativa cloned into the vector pUC57 and inserted into plasmids pYES2 and pGAP using Escherichia coli DH5 alpha strain. The P. pastoris strain X-33 and the S. cerevisiae strain InvSC1 were used for extracellular expression of CHO. After transformation of P. pastoris X-33 with CHO-pGAP construct none of the colonies showed CHO activity. Two samples displayed a band which did not exist in the sample with the empty vector similar to the molecular weight of CHO. The S. cerevisiae strain InvSC1 has been also transformed with CHO-pYES constructs. Three colonies grew on the plate with cells transformed with the construct. One of the samples showed a band corresponding to about 110 kDa, but no CHO activity was recorded in this case either. Cloning of the foreign genes and heterologous expression in yeasts is widely used in biotechnology, but sometimes can be very dependent on the gene sequence and strain used. In order to obtain active CHO enzyme the further studies on purification and refolding of expressed protein are necessary.
AB  - Ugljeni hidrat-oksidaza (CHO) iz zelene salate (Lactuca sativa) je enzim koji je do danas nedovoljno ispitan. Vrlo se malo zna o njegovoj strukturi i funkciji. CHO pripada velikoj familiji ugljenihidrat-oksidaza, koje oksiduju šećere. Svaki od članova ove velike familije dobio je ime po supstratu koji oksiduje. Oksidaze iz ove familije enzima imaju kako sličnu ulogu tako i sličnu strukutru. Sve ili većina ovih enzima su monomeri, čiji se polipeptidni lanac uvija u dva domena. Jedan od domena vezuje flavinski kofaktor, a drugi domen je supstrat vezujući. Većina njih oksidaciju supstrata vrši po takozvanom ping-pong mehanizmu. Sve oksidaze iz karbohidrat-oksidaza familije, pa među njima i enzim koji je predmet ove studije (CHO), danas su našle veliku primenu u industriji. CHO se može primenjivati kako u medicinskoj djagnostici, konkretno u biosenzorima za određivanje glukoze u krvi, u prehrambenoj industriji, poljoprivredi, proizvodnji hleba, deterdženata i u raznim drugim industrijskim oblastima. Problem sa ovim enzimom, kao i sa ostalim članovima ove familije, jeste niska koncentracija u prirodnim izvorima. Zato su danas razvijene različite metode rekombinantne tehnologije, kojima se dobijaju ovi enzimi. U ovom radu opisano je kloniranje gena za CHO iz zelene salate u dve vrste kvasaca (Saccharomyces cerevisiae i Pichia pastoris). Sintetički gen za CHO (1821 bp) iz zelene salate kloniran je u vektor pUC57. Escherichia coli soj DH5α korišćen je za kloniranje gena i održavanje plazmida. P. pastoris soj X-33 i S. cerevisiae soj InvSC1 korišćeni su za ekstracelularnu ekspresiju CHO. Aktivnost CHO određena je ABST esejom, a promena absorbance merena je na 405 nm. Potvrda prisustva enzima rađena je na DNK agaroznoj elektroforezi i SDS-PAGE. Posle transformacije P. pastoris X-33, nijedan od klonova nije pokazivao aktivnost CHO. Posle prve fermentacije, kolonije su testirane na SDS-PAGE. Kako su dva uzorka pokazala trake, koje ne postoje na praznom vektoru, ove trake bi mogle odgovarati željenom enzimu, CHO. Traka se nalazi na molekulskoj masi koja je veća od teoretske (više od 120 kDa). Enzim bi mogao biti glikolizovan i zbog toga pokazivati ovako velike vrednosti za molekulsku masu. S. cerevisiae soj InvSC1 transformisan je konstruktom CHO-pZES. Posle 24 sata, tri kolonije su porasle na ploči na kojoj su bile ćelije transformisane pomenutim konstruktom. Uzorci su testirani na SDS-PAGE. Jedan uzorak je pokazao traku na oko 110 kDa, ali aktivnost CHO nije potvrđena takođe. Cilj ove studije je bio kloniranje CHO u kvascima S. cerevisiae i P. pastoris, kao i njena ekspresija u ovim, danas široko primenjivanim ekspresionim sistemima.
PB  - Assoc Chemical Engineers Serbia, Belgrade
T2  - Hemijska industrija
T1  - Cloning of the gene for a carbohydrate oxidase from Lactuca sativa in the yeasts Saccharomyces cerevisiae and Pichia pastoris
T1  - Kloniranje gena za ugljeni hidrat oksidazu iz biljke Lactuca sativa u kvasce saccharomyces cerevisiae i Pichia pastoris
VL  - 69
IS  - 6
SP  - 689
EP  - 701
DO  - 10.2298/HEMIND140823003T
ER  - 
@article{
author = "Tadić, Vojin and Balaž, Ana Marija and Petric, Marija P. and Milošević, Snežana and Zelenović, Nevena D. and Raspor, Martin Z. and Tadić, Jovan and Prodanović, Radivoje",
year = "2015",
url = "http://cherry.chem.bg.ac.rs/handle/123456789/2040",
abstract = "We have cloned the gene for carbohydrate oxidase (CHO) from Lactuca sativa in two species of yeasts (Saccharomyces cerevisiae and Pichia pastoris). The synthetic gene for the carbohydrate oxidase (1821 bp) from L. sativa cloned into the vector pUC57 and inserted into plasmids pYES2 and pGAP using Escherichia coli DH5 alpha strain. The P. pastoris strain X-33 and the S. cerevisiae strain InvSC1 were used for extracellular expression of CHO. After transformation of P. pastoris X-33 with CHO-pGAP construct none of the colonies showed CHO activity. Two samples displayed a band which did not exist in the sample with the empty vector similar to the molecular weight of CHO. The S. cerevisiae strain InvSC1 has been also transformed with CHO-pYES constructs. Three colonies grew on the plate with cells transformed with the construct. One of the samples showed a band corresponding to about 110 kDa, but no CHO activity was recorded in this case either. Cloning of the foreign genes and heterologous expression in yeasts is widely used in biotechnology, but sometimes can be very dependent on the gene sequence and strain used. In order to obtain active CHO enzyme the further studies on purification and refolding of expressed protein are necessary., Ugljeni hidrat-oksidaza (CHO) iz zelene salate (Lactuca sativa) je enzim koji je do danas nedovoljno ispitan. Vrlo se malo zna o njegovoj strukturi i funkciji. CHO pripada velikoj familiji ugljenihidrat-oksidaza, koje oksiduju šećere. Svaki od članova ove velike familije dobio je ime po supstratu koji oksiduje. Oksidaze iz ove familije enzima imaju kako sličnu ulogu tako i sličnu strukutru. Sve ili većina ovih enzima su monomeri, čiji se polipeptidni lanac uvija u dva domena. Jedan od domena vezuje flavinski kofaktor, a drugi domen je supstrat vezujući. Većina njih oksidaciju supstrata vrši po takozvanom ping-pong mehanizmu. Sve oksidaze iz karbohidrat-oksidaza familije, pa među njima i enzim koji je predmet ove studije (CHO), danas su našle veliku primenu u industriji. CHO se može primenjivati kako u medicinskoj djagnostici, konkretno u biosenzorima za određivanje glukoze u krvi, u prehrambenoj industriji, poljoprivredi, proizvodnji hleba, deterdženata i u raznim drugim industrijskim oblastima. Problem sa ovim enzimom, kao i sa ostalim članovima ove familije, jeste niska koncentracija u prirodnim izvorima. Zato su danas razvijene različite metode rekombinantne tehnologije, kojima se dobijaju ovi enzimi. U ovom radu opisano je kloniranje gena za CHO iz zelene salate u dve vrste kvasaca (Saccharomyces cerevisiae i Pichia pastoris). Sintetički gen za CHO (1821 bp) iz zelene salate kloniran je u vektor pUC57. Escherichia coli soj DH5α korišćen je za kloniranje gena i održavanje plazmida. P. pastoris soj X-33 i S. cerevisiae soj InvSC1 korišćeni su za ekstracelularnu ekspresiju CHO. Aktivnost CHO određena je ABST esejom, a promena absorbance merena je na 405 nm. Potvrda prisustva enzima rađena je na DNK agaroznoj elektroforezi i SDS-PAGE. Posle transformacije P. pastoris X-33, nijedan od klonova nije pokazivao aktivnost CHO. Posle prve fermentacije, kolonije su testirane na SDS-PAGE. Kako su dva uzorka pokazala trake, koje ne postoje na praznom vektoru, ove trake bi mogle odgovarati željenom enzimu, CHO. Traka se nalazi na molekulskoj masi koja je veća od teoretske (više od 120 kDa). Enzim bi mogao biti glikolizovan i zbog toga pokazivati ovako velike vrednosti za molekulsku masu. S. cerevisiae soj InvSC1 transformisan je konstruktom CHO-pZES. Posle 24 sata, tri kolonije su porasle na ploči na kojoj su bile ćelije transformisane pomenutim konstruktom. Uzorci su testirani na SDS-PAGE. Jedan uzorak je pokazao traku na oko 110 kDa, ali aktivnost CHO nije potvrđena takođe. Cilj ove studije je bio kloniranje CHO u kvascima S. cerevisiae i P. pastoris, kao i njena ekspresija u ovim, danas široko primenjivanim ekspresionim sistemima.",
publisher = "Assoc Chemical Engineers Serbia, Belgrade",
journal = "Hemijska industrija",
title = "Cloning of the gene for a carbohydrate oxidase from Lactuca sativa in the yeasts Saccharomyces cerevisiae and Pichia pastoris, Kloniranje gena za ugljeni hidrat oksidazu iz biljke Lactuca sativa u kvasce saccharomyces cerevisiae i Pichia pastoris",
volume = "69",
number = "6",
pages = "689-701",
doi = "10.2298/HEMIND140823003T"
}
Tadić, V., Balaž, A. M., Petric, M. P., Milošević, S., Zelenović, N. D., Raspor, M. Z., Tadić, J.,& Prodanović, R. (2015). Kloniranje gena za ugljeni hidrat oksidazu iz biljke Lactuca sativa u kvasce saccharomyces cerevisiae i Pichia pastoris.
Hemijska industrija
Assoc Chemical Engineers Serbia, Belgrade., 69(6), 689-701.
https://doi.org/10.2298/HEMIND140823003T
Tadić V, Balaž AM, Petric MP, Milošević S, Zelenović ND, Raspor MZ, Tadić J, Prodanović R. Kloniranje gena za ugljeni hidrat oksidazu iz biljke Lactuca sativa u kvasce saccharomyces cerevisiae i Pichia pastoris. Hemijska industrija. 2015;69(6):689-701
Tadić Vojin, Balaž Ana Marija, Petric Marija P., Milošević Snežana, Zelenović Nevena D., Raspor Martin Z., Tadić Jovan, Prodanović Radivoje, "Kloniranje gena za ugljeni hidrat oksidazu iz biljke Lactuca sativa u kvasce saccharomyces cerevisiae i Pichia pastoris" Hemijska industrija, 69, no. 6 (2015):689-701,
https://doi.org/10.2298/HEMIND140823003T .