10.1002/cctc.202001225
ChemCatChem
COMMUNICATION
[9]
J. Golda et al., J. Phys. D: Appl. Phys. 2016, 49, 84003.
Merck, Germany). Absorption was measured at 455 nm and concentration
determined using a calibration curve.
[10] Y. Gorbanev, J. Golda, V. S.-v. d. Gathen, A. Bogaerts, Plasma 2019, 2,
316–327.
[11] a) J. Winter et al., J. Phys. D: Appl. Phys. 2014, 47, 285401; b) Y.
Gorbanev, C. C. W. Verlackt, S. Tinck, E. Tuenter, K. Foubert, P. Cos,
A. Bogaerts, Phys. Chem. Chem. Phys. 2018, 20, 2797–2808;
[12] a) M. Pesic, S. J.-P. Willot, E. Fernández-Fueyo, F. Tieves, M. Alcalde,
F. Hollmann, Z. Naturforsch. C, J. Biosci. 2018; b) W. Zhang et al., Nat.
Catal. 2018, 1, 55; c) Y. Ni, E. Fernández-Fueyo, A. Gomez Baraibar,
R. Ullrich, M. Hofrichter, H. Yanase, M. Alcalde, W. J. H. van Berkel, F.
Hollmann, Angew. Chem. Int. Ed. Engl. 2016, 55, 798–801;
[13] M. Dias Gomes, J. M. Woodley, Molecules 2019, 24, 3573.
[14] a) F. Tieves et al., Angew. Chem. Int. Ed. Engl. 2019; b) W. Zhang, B.
O. Burek, E. Fernández-Fueyo, M. Alcalde, J. Z. Bloh, F. Hollmann,
Angew. Chem. Int. Ed. Engl. 2017, 56, 15451–15455; c) S. J.-P. Willot,
E. Fernández-Fueyo, F. Tieves, M. Pesic, M. Alcalde, I. W. C. E.
Arends, C. B. Park, F. Hollmann, ACS Catal. 2019, 9, 890–894;
[15] a) J. F. Kolb, A.-A. H. Mohamed, R. O. Price, R. J. Swanson, A.
Bowman, R. L. Chiavarini, M. Stacey, K. H. Schoenbach, Appl. Phys.
Lett. 2008, 92, 241501; b) P. Thana, A. Wijaikhum, P. Poramapijitwat,
C. Kuensaen, J. Meerak, A. Ngamjarurojana, S. Sarapirom, D.
Boonyawan, Heliyon 2019, 5, e02455;
Enzyme preparation
In vitro-evolved, recombinant rAaeUPO was purified essentially as
described before.[18] The supernatant of a Pichia pastoris expression
culture was subjected to microfiltration and single-step ion exchange
chromatography, yielding the purified protein. Immobilization was carried
out as described previously.[4] Briefly, HA403 M beads (Resindion, Binasco,
Italy) were washed twice with water and incubated with phosphate buffer
(pH 7) containing 0.5% glutaraldehyde. After 1 h, beads were washed
three times with buffer and enzyme was added at 0.5 nmol per 100 mg
beads. Immobilization was allowed to proceed overnight at 8°C. Binding
efficiency was checked by measuring enzyme activity in the supernatant
and was > 80% in all cases. Roughly 150 mg of protein-loaded beads were
then transferred to a rotating bed reactor that was build in-house by 3D-
printing (dimensions: ⌀ 2 cm x 0.7 cm). The reactor was designed with a
snap-on lid to enable extraction of the enzyme and reuse of the reactor.
The final concentration of rAaeUPO in 5 mL of buffer was approx. 125 nM
(see Tab. S3 for details).
[16] D. Śmiłowicz, F. Kogelheide, K. Stapelmann, P. Awakowicz, N. Metzler-
Nolte, Sci. Rep. 2019, 9, 1–13.
[17] D. Ellerweg, J. Benedikt, A. von Keudell, N. Knake, V. Schulz-von der
Gathen, New J. Phys. 2010, 12, 13021.
[18] P. Molina-Espeja, S. Ma, D. M. Mate, R. Ludwig, M. Alcalde, Enzyme
Microb. Technol. 2015, 73-74, 29–33.
Conversion of ETBE
To generate plasma-treated buffer, 5 mL of buffer were treated with the
µAPPJ for 15 min with constant stirring. Subsequently, 50 mM of ETBE
were added and the reaction solution was mixed for 15 min by overhead
rotation to allow for the substrate to go into solution. Then, 50 nM of
rAaeUPO were added and the solution incubated at 30°C for 15 min with
constant shaking.
When the rotating bed reactor was used, 5 mL of buffer containing 50 mM
of ETBE were mixed and placed into a suitable vessel with the rotating bed
reactor. Plasma exposure was conducted at approx. 4 mm distance
between the nozzle and the liquid surface.
Analysis of (R)-1-Phol
Aliquots of 150 µL were withdrawn and mixed with the same volume of
ethyl acetate containing 2 mM of 1-octanol as internal standard. The
organic phase was transferred to a new vial, dried with MgSO4, and
subjected to gas chromatography. Samples were analyzed using a
Shimadzu 2010 system with a Hydrodex β-6TBDM column (Macherey-
Nagel, Germany) in an isothermal program (125°C, 10 min).
Concentrations were determined by a calibration curve with racemic 1-
Phol.
Acknowledgements
We thank Marco Krewing for fruitful discussions and the workshop
of the Department of Biology and Biotechnology (Bochum) for
constructing the rotating bed reactor. Funding was provided by
CRC1316-1 to JEB and PA and RTG2341 to JEB.
Keywords: atmospheric pressure plasma • biocatalysis •
hydrogen peroxide • peroxygenase • plasma chemistry
[1]
Y. Wang, D. Lan, R. Durrani, F. Hollmann, Curr. Opin. Chem. Biol.
2017, 37, 1–9.
[2]
[3]
B. O. Burek et al., ChemCatChem 2019.
S. Bormann, A. Gomez Baraibar, Y. Ni, D. Holtmann, F. Hollmann,
Catal. Sci. Technol. 2015, 5, 2038–2052.
[4]
A. Yayci, Á. Gómez Baraibar, M. Krewing, E. Fernandez Fueyo, F.
Hollmann, M. Alcalde, R. Kourist, J. E. Bandow, ChemSusChem 2020,
13, 2072–2079.
[5]
[6]
[7]
R. Brandenburg, Plasma Sources Sci. Technol. 2017, 26, 53001.
P. J. Bruggeman et al., Plasma Sources Sci. Technol. 2016, 25, 53002.
F. Kogelheide, B. Offerhaus, N. Bibinov, P. Krajinski, L. Schücke, J.
Schulze, K. Stapelmann, P. Awakowicz, Plasma. Process. Polym. 2020,
17, 1900126.
[8]
M. Kuchenbecker, N. Bibinov, A. Kaemlimg, D. Wandke, P. Awakowicz,
W. Viöl, J. Phys. D: Appl. Phys. 2009, 42, 45212.
5
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