Preprint Article Version 1 Preserved in Portico This version is not peer-reviewed

VpStyA1 and VpStyA2B of Variovorax paradoxus EPS: Rather an Aryl Alkyl Sulfoxidase than a Styrene Epoxidizing Monooxygenase

Dirk Tischler
* ORCID logo ,
Ringo Schwabe
,
Lucas Siegel
,
Kristin Joffroy
,
Stefan R. Kaschabek
,
Anika Scholtissek
,
Thomas Heine
ORCID logo
Version 1 : Received: 25 January 2018 / Approved: 25 January 2018 / Online: 25 January 2018 (17:14:33 CET)

How to cite: Tischler, D.; Schwabe, R.; Siegel, L.; Joffroy, K.; Kaschabek, S.R.; Scholtissek, A.; Heine, T. VpStyA1 and VpStyA2B of Variovorax paradoxus EPS: Rather an Aryl Alkyl Sulfoxidase than a Styrene Epoxidizing Monooxygenase. Preprints 2018, 2018010245. https://doi.org/10.20944/preprints201801.0245.v1 Tischler, D.; Schwabe, R.; Siegel, L.; Joffroy, K.; Kaschabek, S.R.; Scholtissek, A.; Heine, T. VpStyA1 and VpStyA2B of Variovorax paradoxus EPS: Rather an Aryl Alkyl Sulfoxidase than a Styrene Epoxidizing Monooxygenase. Preprints 2018, 2018010245. https://doi.org/10.20944/preprints201801.0245.v1

Abstract

VpStyA1 and VpStyA2B of Variovorax paradoxus EPS is annotated and characterized as the first representative of an E2-type styrene monooxygenase of proteobacteria. It comprises a single epoxidase (VpStyA1) and a fusion protein (VpStyA2B) which serves mainly as NADH:FAD-oxidoreductase. VpStyA2B had a Km of 33.6 ± 4.0 µM for FAD and a kcat of 22.3 ± 1.1 s-1. VpStyA2B and VpStyA1 showed monooxygenase activity on styrene of 0.14 U mg-1 and 0.46 U mg-1 as well as on benzyl methyl sulfide of 1.62 U mg-1 and of 3.11 U mg-1. A putative fusion region at position 408 (AREAV) was mutated to provide insights on VpStyA2B-function. The best mutant (408-AAAAA) obtained showed a 6.6-times higher affinity for FAD while keeping the NADH-affinity and -oxidation activity. Corresponding epoxidase activity increased (1.6-times). But, other mutants showed still NADH:FAD-oxidoreductase activity, but lost mostly their epoxidase activity indicating effects on the monooxygenase-part as well. Thus, this monooxygenase system represents an interesting candidate for biocatalyst development.

Keywords

sulfoxidation; epoxidation; two-component monooxygenase; flavoprotein; enantioselective biotransformation; fusion protein; protein linker; soil microorganism

Subject

Biology and Life Sciences, Biochemistry and Molecular Biology

Copyright: This is an open access article distributed under the Creative Commons Attribution License which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

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