The aim of this research was to improve the operational stability and enable the reusability of ω-transaminase for synthesis of new enantiopure chiral amines of steroids. Dihydrotestosterone was used to optimize the synthetic procedure of corresponding amino-steroid on a larger scale. The obtained product 3α-amino-5α-androstan-17β-ol was isolated and characterized. The enzyme was immobilized on a methacrylate-based carrier, giving the specific activity of 1.84 U/g of dry polymer. Higher residual activity of the immobilized enzyme in comparison to the soluble form (100 % versus 35%) after 24 h incubation in 35 % dimethylformamide (DMF) was obtained. The soluble enzyme retained 19 % of the initial activity after 2 h incubation in 35 % DMF at 70 °C, while the activity of the immobilized enzyme decreased only to 75 %. Immobilized retained 85 % of initial activity after ten consecutive cycles of 3α-amino-5α-androstan-17β-ol synthesis. We have tested the specificity of the ArRMut11 variant, ...further increased its stability by immobilization, and used it in several cycles for the synthesis of 3α-amino-5α-androstan-17β-ol. We showed that the enzyme previously evolved for higher stability as the immobilized variant showed more increased stability and high reusability that can more effectively be applied for the biosynthesis of amino steroids.
TY - JOUR
AU - Kaličanin, Nevena
AU - Kovačević, Gordana
AU - Spasojević, Milica
AU - Prodanović, Olivera
AU - Jovanović-Šanta, Suzana
AU - Škorić, Dušan
AU - Opsenica, Dejan
AU - Prodanović, Radivoje
PY - 2022
UR - http://rimsi.imsi.bg.ac.rs/handle/123456789/1768
AB - The aim of this research was to improve the operational stability and enable the reusability of ω-transaminase for synthesis of new enantiopure chiral amines of steroids. Dihydrotestosterone was used to optimize the synthetic procedure of corresponding amino-steroid on a larger scale. The obtained product 3α-amino-5α-androstan-17β-ol was isolated and characterized. The enzyme was immobilized on a methacrylate-based carrier, giving the specific activity of 1.84 U/g of dry polymer. Higher residual activity of the immobilized enzyme in comparison to the soluble form (100 % versus 35%) after 24 h incubation in 35 % dimethylformamide (DMF) was obtained. The soluble enzyme retained 19 % of the initial activity after 2 h incubation in 35 % DMF at 70 °C, while the activity of the immobilized enzyme decreased only to 75 %. Immobilized retained 85 % of initial activity after ten consecutive cycles of 3α-amino-5α-androstan-17β-ol synthesis. We have tested the specificity of the ArRMut11 variant, further increased its stability by immobilization, and used it in several cycles for the synthesis of 3α-amino-5α-androstan-17β-ol. We showed that the enzyme previously evolved for higher stability as the immobilized variant showed more increased stability and high reusability that can more effectively be applied for the biosynthesis of amino steroids.
PB - Elsevier
T2 - Process Biochemistry
T1 - Immobilization of ArRMut11 omega-transaminase for increased operational stability and reusability in the synthesis of 3α-amino-5α-androstan-17β-ol
EP - 680
SP - 674
VL - 121
DO - 10.1016/j.procbio.2022.08.016
ER -
@article{
author = "Kaličanin, Nevena and Kovačević, Gordana and Spasojević, Milica and Prodanović, Olivera and Jovanović-Šanta, Suzana and Škorić, Dušan and Opsenica, Dejan and Prodanović, Radivoje",
year = "2022",
abstract = "The aim of this research was to improve the operational stability and enable the reusability of ω-transaminase for synthesis of new enantiopure chiral amines of steroids. Dihydrotestosterone was used to optimize the synthetic procedure of corresponding amino-steroid on a larger scale. The obtained product 3α-amino-5α-androstan-17β-ol was isolated and characterized. The enzyme was immobilized on a methacrylate-based carrier, giving the specific activity of 1.84 U/g of dry polymer. Higher residual activity of the immobilized enzyme in comparison to the soluble form (100 % versus 35%) after 24 h incubation in 35 % dimethylformamide (DMF) was obtained. The soluble enzyme retained 19 % of the initial activity after 2 h incubation in 35 % DMF at 70 °C, while the activity of the immobilized enzyme decreased only to 75 %. Immobilized retained 85 % of initial activity after ten consecutive cycles of 3α-amino-5α-androstan-17β-ol synthesis. We have tested the specificity of the ArRMut11 variant, further increased its stability by immobilization, and used it in several cycles for the synthesis of 3α-amino-5α-androstan-17β-ol. We showed that the enzyme previously evolved for higher stability as the immobilized variant showed more increased stability and high reusability that can more effectively be applied for the biosynthesis of amino steroids.",
publisher = "Elsevier",
journal = "Process Biochemistry",
title = "Immobilization of ArRMut11 omega-transaminase for increased operational stability and reusability in the synthesis of 3α-amino-5α-androstan-17β-ol",
pages = "680-674",
volume = "121",
doi = "10.1016/j.procbio.2022.08.016"
}
Kaličanin, N., Kovačević, G., Spasojević, M., Prodanović, O., Jovanović-Šanta, S., Škorić, D., Opsenica, D.,& Prodanović, R.. (2022). Immobilization of ArRMut11 omega-transaminase for increased operational stability and reusability in the synthesis of 3α-amino-5α-androstan-17β-ol. in Process Biochemistry
Elsevier., 121, 674-680.
https://doi.org/10.1016/j.procbio.2022.08.016
Kaličanin N, Kovačević G, Spasojević M, Prodanović O, Jovanović-Šanta S, Škorić D, Opsenica D, Prodanović R. Immobilization of ArRMut11 omega-transaminase for increased operational stability and reusability in the synthesis of 3α-amino-5α-androstan-17β-ol. in Process Biochemistry. 2022;121:674-680.
doi:10.1016/j.procbio.2022.08.016 .
Kaličanin, Nevena, Kovačević, Gordana, Spasojević, Milica, Prodanović, Olivera, Jovanović-Šanta, Suzana, Škorić, Dušan, Opsenica, Dejan, Prodanović, Radivoje, "Immobilization of ArRMut11 omega-transaminase for increased operational stability and reusability in the synthesis of 3α-amino-5α-androstan-17β-ol" in Process Biochemistry, 121 (2022):674-680,
https://doi.org/10.1016/j.procbio.2022.08.016 . .
