Details
| Original language | English |
|---|---|
| Pages (from-to) | 11-17 |
| Number of pages | 7 |
| Journal | Enzyme and microbial technology |
| Volume | 106 |
| Early online date | 3 Jul 2017 |
| Publication status | Published - Nov 2017 |
| Externally published | Yes |
Abstract
FAD-dependent Baeyer-Villiger monooxygenases (BVMOs) have proven to be useful biocatalysts in the selective and specific oxygenation of various ketones. Despite the cloning, heterologous expression and characterization of close to 80 members of this enzyme family, some sub-groups of BVMOs still remain underrepresented and their evolutionary relationship uncertain. Until recently, very few fungal BVMOs have been described. Our previous investigations into BVMOs from the fungus Aspergillus flavus, yielded very little activity on simple cyclic ketones. Here we report on another four BVMOs from A. flavus that are more closely related to cyclohexanone monooxygenase (CHMO) from Acinetobacter sp. NCIMB 9871. Evolutionary analysis with other characterized BVMOs show their closest relationship to be with either cycloalkanone monooxygenase (CAMO) or 2-oxo-Δ3-4,5,5-trimethylcyclopentenylacetyl-coenzyme A monooxygenase (OTEMO). The OTEMO-related BVMOAFL706 and BVMOAFL334 were heterologously expressed in E. coli, purified and shown to be able to convert a range of cyclic and substituted cyclic ketones. Of the unsubstituted cyclic ketones, cyclohexanone showed the highest conversion with maximum turnover frequencies reaching 4.3 s−1 for BVMOAFL706. Unlike CHMOacinet, and many of the closely related BVMOs, no substrate inhibition was observed with cyclohexanone to a concentration of up to 30 mM, creating the possibility for applications requiring high substrate loading. Aliphatic ketones were also readily converted with excellent regioselectivity. Similar to CHMOacinet, acetophenones were not converted and the oxidation of rac-cis-bicyclo[3.2.0]hept-2-en-6-one occurs enantiodivergently, with the (1R,5S) isomer converted to the “normal” lactone and the (1S,5R) isomer to the “abnormal” lactone.
Keywords
- Aspergillus flavus, Baeyer-Villiger monooxygenase, Lactone
ASJC Scopus subject areas
- Biochemistry, Genetics and Molecular Biology(all)
- Biotechnology
- Chemical Engineering(all)
- Bioengineering
- Biochemistry, Genetics and Molecular Biology(all)
- Biochemistry
- Immunology and Microbiology(all)
- Applied Microbiology and Biotechnology
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In: Enzyme and microbial technology, Vol. 106, 11.2017, p. 11-17.
Research output: Contribution to journal › Article › Research › peer review
}
TY - JOUR
T1 - Fungal BVMOs as alternatives to cyclohexanone monooxygenase
AU - Mthethwa, Katlego Siphamandla
AU - Kassier, Karin
AU - Engel, Jennifer
AU - Kara, Selin
AU - Smit, Martha Sophia
AU - Opperman, Diederik Johannes
N1 - Funding Information: The authors would like to thank Mr. Sarel Marais for GC-(MS) analyses. This study was funded by the National Research Foundation (NRF), South Africa (Grant No. TTK13060518534) and the Deutsche Forschungsgemeinschaft (DFG), Germany(Grant No. KA 4399/1-1).
PY - 2017/11
Y1 - 2017/11
N2 - FAD-dependent Baeyer-Villiger monooxygenases (BVMOs) have proven to be useful biocatalysts in the selective and specific oxygenation of various ketones. Despite the cloning, heterologous expression and characterization of close to 80 members of this enzyme family, some sub-groups of BVMOs still remain underrepresented and their evolutionary relationship uncertain. Until recently, very few fungal BVMOs have been described. Our previous investigations into BVMOs from the fungus Aspergillus flavus, yielded very little activity on simple cyclic ketones. Here we report on another four BVMOs from A. flavus that are more closely related to cyclohexanone monooxygenase (CHMO) from Acinetobacter sp. NCIMB 9871. Evolutionary analysis with other characterized BVMOs show their closest relationship to be with either cycloalkanone monooxygenase (CAMO) or 2-oxo-Δ3-4,5,5-trimethylcyclopentenylacetyl-coenzyme A monooxygenase (OTEMO). The OTEMO-related BVMOAFL706 and BVMOAFL334 were heterologously expressed in E. coli, purified and shown to be able to convert a range of cyclic and substituted cyclic ketones. Of the unsubstituted cyclic ketones, cyclohexanone showed the highest conversion with maximum turnover frequencies reaching 4.3 s−1 for BVMOAFL706. Unlike CHMOacinet, and many of the closely related BVMOs, no substrate inhibition was observed with cyclohexanone to a concentration of up to 30 mM, creating the possibility for applications requiring high substrate loading. Aliphatic ketones were also readily converted with excellent regioselectivity. Similar to CHMOacinet, acetophenones were not converted and the oxidation of rac-cis-bicyclo[3.2.0]hept-2-en-6-one occurs enantiodivergently, with the (1R,5S) isomer converted to the “normal” lactone and the (1S,5R) isomer to the “abnormal” lactone.
AB - FAD-dependent Baeyer-Villiger monooxygenases (BVMOs) have proven to be useful biocatalysts in the selective and specific oxygenation of various ketones. Despite the cloning, heterologous expression and characterization of close to 80 members of this enzyme family, some sub-groups of BVMOs still remain underrepresented and their evolutionary relationship uncertain. Until recently, very few fungal BVMOs have been described. Our previous investigations into BVMOs from the fungus Aspergillus flavus, yielded very little activity on simple cyclic ketones. Here we report on another four BVMOs from A. flavus that are more closely related to cyclohexanone monooxygenase (CHMO) from Acinetobacter sp. NCIMB 9871. Evolutionary analysis with other characterized BVMOs show their closest relationship to be with either cycloalkanone monooxygenase (CAMO) or 2-oxo-Δ3-4,5,5-trimethylcyclopentenylacetyl-coenzyme A monooxygenase (OTEMO). The OTEMO-related BVMOAFL706 and BVMOAFL334 were heterologously expressed in E. coli, purified and shown to be able to convert a range of cyclic and substituted cyclic ketones. Of the unsubstituted cyclic ketones, cyclohexanone showed the highest conversion with maximum turnover frequencies reaching 4.3 s−1 for BVMOAFL706. Unlike CHMOacinet, and many of the closely related BVMOs, no substrate inhibition was observed with cyclohexanone to a concentration of up to 30 mM, creating the possibility for applications requiring high substrate loading. Aliphatic ketones were also readily converted with excellent regioselectivity. Similar to CHMOacinet, acetophenones were not converted and the oxidation of rac-cis-bicyclo[3.2.0]hept-2-en-6-one occurs enantiodivergently, with the (1R,5S) isomer converted to the “normal” lactone and the (1S,5R) isomer to the “abnormal” lactone.
KW - Aspergillus flavus
KW - Baeyer-Villiger monooxygenase
KW - Lactone
UR - http://www.scopus.com/inward/record.url?scp=85021891277&partnerID=8YFLogxK
U2 - 10.1016/j.enzmictec.2017.06.017
DO - 10.1016/j.enzmictec.2017.06.017
M3 - Article
C2 - 28859804
AN - SCOPUS:85021891277
VL - 106
SP - 11
EP - 17
JO - Enzyme and microbial technology
JF - Enzyme and microbial technology
SN - 0141-0229
ER -