TY  - JOUR
AU  - Gursky, Lucas J.
AU  - Nikodinović-Runić, Jasmina
AU  - Feenstra, K. Anton
AU  - O'Connor, Kevin E.
PY  - 2010
UR  - https://cherry.chem.bg.ac.rs/handle/123456789/1041
AB  - The styAB genes from Pseudomonas putida CA-3, which encode styrene monooxygenase, were subjected to three rounds of in vitro evolution using error-prone polymerase chain reaction with a view to improving the rate of styrene oxide and indene oxide formation. Improvements in styrene monooxygenase activity were monitored using an indole to indigo conversion assay. Each round of random mutagenesis generated variants improved in indigo formation with third round variants improved nine- to 12-fold over the wild type enzyme. Each round of in vitro evolution resulted in two to three amino acid substitutions in styrene monooxygenase. While the majority of mutations occurred in styA (oxygenase), mutations were also observed in styB (reductase). A mutation resulting in the substitution of valine with isoleucine at amino acid residue 303 occurred near the styrene and flavin adenine dinucleotide binding site of styrene monooxygenase. One mutation caused a shift in the reading frame in styA and resulted in a StyA variant that is 19 amino acids longer than the wild-type protein. Whole cells expressing the best styrene monooxygenase variants (round 3) exhibited eight- and 12-fold improvements in styrene and indene oxidation rates compared to the wild-type enzyme. In all cases, a single enantiomer, (S)-styrene oxide, was formed from styrene while (1S,2R)-indene oxide was the predominant enantiomer (e.e. 97%) formed from indene. The average yield of styrene oxide and indene oxide from their respective alkene substrates was 65% and 90%, respectively.
PB  - Springer, New York
T2  - Applied Microbiology and Biotechnology
T1  - In vitro evolution of styrene monooxygenase from Pseudomonas putida CA-3 for improved epoxide synthesis
VL  - 85
IS  - 4
SP  - 995
EP  - 1004
DO  - 10.1007/s00253-009-2096-3
ER  - 

@article{
author = "Gursky, Lucas J. and Nikodinović-Runić, Jasmina and Feenstra, K. Anton and O'Connor, Kevin E.",
year = "2010",
abstract = "The styAB genes from Pseudomonas putida CA-3, which encode styrene monooxygenase, were subjected to three rounds of in vitro evolution using error-prone polymerase chain reaction with a view to improving the rate of styrene oxide and indene oxide formation. Improvements in styrene monooxygenase activity were monitored using an indole to indigo conversion assay. Each round of random mutagenesis generated variants improved in indigo formation with third round variants improved nine- to 12-fold over the wild type enzyme. Each round of in vitro evolution resulted in two to three amino acid substitutions in styrene monooxygenase. While the majority of mutations occurred in styA (oxygenase), mutations were also observed in styB (reductase). A mutation resulting in the substitution of valine with isoleucine at amino acid residue 303 occurred near the styrene and flavin adenine dinucleotide binding site of styrene monooxygenase. One mutation caused a shift in the reading frame in styA and resulted in a StyA variant that is 19 amino acids longer than the wild-type protein. Whole cells expressing the best styrene monooxygenase variants (round 3) exhibited eight- and 12-fold improvements in styrene and indene oxidation rates compared to the wild-type enzyme. In all cases, a single enantiomer, (S)-styrene oxide, was formed from styrene while (1S,2R)-indene oxide was the predominant enantiomer (e.e. 97%) formed from indene. The average yield of styrene oxide and indene oxide from their respective alkene substrates was 65% and 90%, respectively.",
publisher = "Springer, New York",
journal = "Applied Microbiology and Biotechnology",
title = "In vitro evolution of styrene monooxygenase from Pseudomonas putida CA-3 for improved epoxide synthesis",
volume = "85",
number = "4",
pages = "995-1004",
doi = "10.1007/s00253-009-2096-3"
}

Gursky, L. J., Nikodinović-Runić, J., Feenstra, K. A.,& O'Connor, K. E.. (2010). In vitro evolution of styrene monooxygenase from Pseudomonas putida CA-3 for improved epoxide synthesis. in Applied Microbiology and Biotechnology
Springer, New York., 85(4), 995-1004.
https://doi.org/10.1007/s00253-009-2096-3

Gursky LJ, Nikodinović-Runić J, Feenstra KA, O'Connor KE. In vitro evolution of styrene monooxygenase from Pseudomonas putida CA-3 for improved epoxide synthesis. in Applied Microbiology and Biotechnology. 2010;85(4):995-1004.
doi:10.1007/s00253-009-2096-3 .

Gursky, Lucas J., Nikodinović-Runić, Jasmina, Feenstra, K. Anton, O'Connor, Kevin E., "In vitro evolution of styrene monooxygenase from Pseudomonas putida CA-3 for improved epoxide synthesis" in Applied Microbiology and Biotechnology, 85, no. 4 (2010):995-1004,
https://doi.org/10.1007/s00253-009-2096-3 . .