Importance of N-terminal proline for the promiscuous activity of 4-oxalocrotonate tautomerase (4-OT)
2016
Authors
Lazić, JelenaSpasić, Jelena
Francuski, Djordje
Tokić-Vujosević, Zorana
Nikodinović-Runić, Jasmina
Maslak, Veselin
Đokić, Lidija
Article (Published version)
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Michael addition of aldehydes to nitro-olefins is an effective method to obtain useful chiral gamma-nitroaldehydes. gamma-Nitroaldehydes are precursors for chiral gamma-aminobutyric acid analogues, which have numerous pharmacological activities and are used for the treatment of neurological disorders. A whole-cell system based on recombinantly expressed 4-oxalocrotonate tautomerase (4-OT) was developed and shown to be an effective biocatalyst for the Michael addition of branched aldehydes to beta-nitrostyrenes. The aim of this study was to investigate the influence of the substitution of the N-terminal proline with lysine and arginine, both containing a reactive epsilon-amino group, on the Michael addition catalyzed by 4-OT. First, the effects of these mutations were examined by in silico analysis, followed by the generation of three terminal lysine mutants. The generated mutants, 4-OT_K, 4-OT_PK and 4-OT_KK were tested for their ability to utilise beta-nitrostyrene (1), (E)-1-nitro-2-...(2-thienyl)ethene (2) and trans-p-chloro-beta-nitrostyrene (3) as Michael acceptors with isobutanal (2-methylpropanal) as the donor. For comparison, the lithium salt of lysine was used in the same organocatalytic reactions. In general, the introduction of lysine had a negative effect on Michael additions based on overall product yields. However, additional lysine residues at the N-terminus of the protein resulted in structural changes that enhanced the activity towards 2 and 3. Therefore, the N-terminal proline is important for 4-OT-catalysed Michael-additions, but it is not essential.
Keywords:
organocatalysis / mutagenesis / lysine / biocatalysis / 4-oxalocrotonate tautomeraseSource:
Journal of the Serbian Chemical Society, 2016, 81, 8, 871-881Publisher:
- Srpsko hemijsko društvo, Beograd
Funding / projects:
- Microbial diversity study and characterization of beneficial environmental microorganisms (RS-MESTD-Basic Research (BR or ON)-173048)
- Design, synthesis and investigations of fullerene based nanomolecular machines (RS-MESTD-Basic Research (BR or ON)-172002)
DOI: 10.2298/JSC160222053L
ISSN: 0352-5139
WoS: 000384930600003
Scopus: 2-s2.0-84987792424
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Institution/Community
Institut za molekularnu genetiku i genetičko inženjerstvoTY - JOUR AU - Lazić, Jelena AU - Spasić, Jelena AU - Francuski, Djordje AU - Tokić-Vujosević, Zorana AU - Nikodinović-Runić, Jasmina AU - Maslak, Veselin AU - Đokić, Lidija PY - 2016 UR - https://imagine.imgge.bg.ac.rs/handle/123456789/929 AB - Michael addition of aldehydes to nitro-olefins is an effective method to obtain useful chiral gamma-nitroaldehydes. gamma-Nitroaldehydes are precursors for chiral gamma-aminobutyric acid analogues, which have numerous pharmacological activities and are used for the treatment of neurological disorders. A whole-cell system based on recombinantly expressed 4-oxalocrotonate tautomerase (4-OT) was developed and shown to be an effective biocatalyst for the Michael addition of branched aldehydes to beta-nitrostyrenes. The aim of this study was to investigate the influence of the substitution of the N-terminal proline with lysine and arginine, both containing a reactive epsilon-amino group, on the Michael addition catalyzed by 4-OT. First, the effects of these mutations were examined by in silico analysis, followed by the generation of three terminal lysine mutants. The generated mutants, 4-OT_K, 4-OT_PK and 4-OT_KK were tested for their ability to utilise beta-nitrostyrene (1), (E)-1-nitro-2-(2-thienyl)ethene (2) and trans-p-chloro-beta-nitrostyrene (3) as Michael acceptors with isobutanal (2-methylpropanal) as the donor. For comparison, the lithium salt of lysine was used in the same organocatalytic reactions. In general, the introduction of lysine had a negative effect on Michael additions based on overall product yields. However, additional lysine residues at the N-terminus of the protein resulted in structural changes that enhanced the activity towards 2 and 3. Therefore, the N-terminal proline is important for 4-OT-catalysed Michael-additions, but it is not essential. PB - Srpsko hemijsko društvo, Beograd T2 - Journal of the Serbian Chemical Society T1 - Importance of N-terminal proline for the promiscuous activity of 4-oxalocrotonate tautomerase (4-OT) EP - 881 IS - 8 SP - 871 VL - 81 DO - 10.2298/JSC160222053L ER -
@article{ author = "Lazić, Jelena and Spasić, Jelena and Francuski, Djordje and Tokić-Vujosević, Zorana and Nikodinović-Runić, Jasmina and Maslak, Veselin and Đokić, Lidija", year = "2016", abstract = "Michael addition of aldehydes to nitro-olefins is an effective method to obtain useful chiral gamma-nitroaldehydes. gamma-Nitroaldehydes are precursors for chiral gamma-aminobutyric acid analogues, which have numerous pharmacological activities and are used for the treatment of neurological disorders. A whole-cell system based on recombinantly expressed 4-oxalocrotonate tautomerase (4-OT) was developed and shown to be an effective biocatalyst for the Michael addition of branched aldehydes to beta-nitrostyrenes. The aim of this study was to investigate the influence of the substitution of the N-terminal proline with lysine and arginine, both containing a reactive epsilon-amino group, on the Michael addition catalyzed by 4-OT. First, the effects of these mutations were examined by in silico analysis, followed by the generation of three terminal lysine mutants. The generated mutants, 4-OT_K, 4-OT_PK and 4-OT_KK were tested for their ability to utilise beta-nitrostyrene (1), (E)-1-nitro-2-(2-thienyl)ethene (2) and trans-p-chloro-beta-nitrostyrene (3) as Michael acceptors with isobutanal (2-methylpropanal) as the donor. For comparison, the lithium salt of lysine was used in the same organocatalytic reactions. In general, the introduction of lysine had a negative effect on Michael additions based on overall product yields. However, additional lysine residues at the N-terminus of the protein resulted in structural changes that enhanced the activity towards 2 and 3. Therefore, the N-terminal proline is important for 4-OT-catalysed Michael-additions, but it is not essential.", publisher = "Srpsko hemijsko društvo, Beograd", journal = "Journal of the Serbian Chemical Society", title = "Importance of N-terminal proline for the promiscuous activity of 4-oxalocrotonate tautomerase (4-OT)", pages = "881-871", number = "8", volume = "81", doi = "10.2298/JSC160222053L" }
Lazić, J., Spasić, J., Francuski, D., Tokić-Vujosević, Z., Nikodinović-Runić, J., Maslak, V.,& Đokić, L.. (2016). Importance of N-terminal proline for the promiscuous activity of 4-oxalocrotonate tautomerase (4-OT). in Journal of the Serbian Chemical Society Srpsko hemijsko društvo, Beograd., 81(8), 871-881. https://doi.org/10.2298/JSC160222053L
Lazić J, Spasić J, Francuski D, Tokić-Vujosević Z, Nikodinović-Runić J, Maslak V, Đokić L. Importance of N-terminal proline for the promiscuous activity of 4-oxalocrotonate tautomerase (4-OT). in Journal of the Serbian Chemical Society. 2016;81(8):871-881. doi:10.2298/JSC160222053L .
Lazić, Jelena, Spasić, Jelena, Francuski, Djordje, Tokić-Vujosević, Zorana, Nikodinović-Runić, Jasmina, Maslak, Veselin, Đokić, Lidija, "Importance of N-terminal proline for the promiscuous activity of 4-oxalocrotonate tautomerase (4-OT)" in Journal of the Serbian Chemical Society, 81, no. 8 (2016):871-881, https://doi.org/10.2298/JSC160222053L . .