TY - JOUR
T1 - Engineering cyanobacterial chassis for improved electron supply toward a heterologous ene-reductase
AU - Spasic, Jelena
AU - Oliveira, Paulo
AU - Pacheco, Catarina
AU - Kourist, Robert
AU - Tamagnini, Paula
N1 - Funding Information:
This project has received funding from the European Union’s Horizon 2020 research and innovation program under the Marie Skłodowska-Curie grant agreement No 764920 . CCP and PO acknowledge the Fundação para a Ciência e a Tecnologia ( FCT )/ Ministério da Ciência, Tecnologia e Ensino Superior for the Assistant Researcher contract CEECIND/00259/2017 , and for the FCT Investigator Grant IF/00256/2015 , respectively. The authors acknowledge the support of the i3S Scientific Platform BioSciences Screening member of the national infrastructure PT-OPENSCREEN ( NORTE-01-0145-FEDER-085468 ).
Funding Information:
This project has received funding from the European Union's Horizon 2020 research and innovation program under the Marie Skłodowska-Curie grant agreement No 764920. CCP and PO acknowledge the Fundação para a Ciência e a Tecnologia (FCT)/Ministério da Ciência, Tecnologia e Ensino Superior for the Assistant Researcher contract CEECIND/00259/2017, and for the FCT Investigator Grant IF/00256/2015, respectively. The authors acknowledge the support of the i3S Scientific Platform BioSciences Screening member of the national infrastructure PT-OPENSCREEN (NORTE-01-0145-FEDER-085468).
Publisher Copyright:
© 2022 Elsevier B.V.
PY - 2022/12/10
Y1 - 2022/12/10
N2 - Cyanobacteria are noteworthy hosts for industrially relevant redox reactions, owing to a light-driven cofactor recycling system using water as electron donor. Customizing Synechocystis sp. PCC 6803 chassis by redirecting electron flow offers a particularly interesting approach to further improve light-driven biotransformations. Therefore, different chassis expressing the heterologous ene-reductase YqjM (namely ΔhoxYH, Δflv3, ΔndhD2 and ΔhoxYHΔflv3) were generated/evaluated. The results showed the robustness of the chassis, that exhibited growth and oxygen evolution rates similar to Synechocystis wild-type, even when expressing YqjM. By engineering the electron flow, the YqjM light-driven stereoselective reduction of 2-methylmaleimide to 2-methylsuccinimide was significantly enhanced in all chassis. In the best performing chassis (ΔhoxYH, lacking an active bidirectional hydrogenase) a 39 % increase was observed, reaching an in vivo specific activity of 116 U gDCW−1 and an initial reaction rate of 16.7 mM h−1. In addition, the presence of the heterologous YqjM mitigated substrate toxicity, and the conversion of 2-methylmaleimide increased oxygen evolution rates, in particular at higher light intensity. In conclusion, this work demonstrates that rational engineering of electron transfer pathways is a valid strategy to increase in vivo specific activities and initial reaction rates in cyanobacterial chassis harboring oxidoreductases.
AB - Cyanobacteria are noteworthy hosts for industrially relevant redox reactions, owing to a light-driven cofactor recycling system using water as electron donor. Customizing Synechocystis sp. PCC 6803 chassis by redirecting electron flow offers a particularly interesting approach to further improve light-driven biotransformations. Therefore, different chassis expressing the heterologous ene-reductase YqjM (namely ΔhoxYH, Δflv3, ΔndhD2 and ΔhoxYHΔflv3) were generated/evaluated. The results showed the robustness of the chassis, that exhibited growth and oxygen evolution rates similar to Synechocystis wild-type, even when expressing YqjM. By engineering the electron flow, the YqjM light-driven stereoselective reduction of 2-methylmaleimide to 2-methylsuccinimide was significantly enhanced in all chassis. In the best performing chassis (ΔhoxYH, lacking an active bidirectional hydrogenase) a 39 % increase was observed, reaching an in vivo specific activity of 116 U gDCW−1 and an initial reaction rate of 16.7 mM h−1. In addition, the presence of the heterologous YqjM mitigated substrate toxicity, and the conversion of 2-methylmaleimide increased oxygen evolution rates, in particular at higher light intensity. In conclusion, this work demonstrates that rational engineering of electron transfer pathways is a valid strategy to increase in vivo specific activities and initial reaction rates in cyanobacterial chassis harboring oxidoreductases.
KW - Biocatalysis
KW - Biotransformation
KW - Cyanobacteria
KW - Synechocystis
KW - Synthetic biology
UR - http://www.scopus.com/inward/record.url?scp=85142136798&partnerID=8YFLogxK
U2 - 10.1016/j.jbiotec.2022.11.005
DO - 10.1016/j.jbiotec.2022.11.005
M3 - Article
C2 - 36370921
AN - SCOPUS:85142136798
SN - 0168-1656
VL - 360
SP - 152
EP - 159
JO - Journal of Biotechnology
JF - Journal of Biotechnology
ER -