Enhanced Production of D-Lactate in Cyanobacteria by Re-Routing Photosynthetic Cyclic and Pseudo-Cyclic Electron Flow

Abstract

Cyanobacteria are promising chassis strains for the photosynthetic production of platform and specialty chemicals from carbon dioxide. Their efficient light harvesting and metabolic flexibility abilities have allowed a wide range of biomolecules, such as the bioplastic polylactate precursor D-lactate, to be produced, though usually at relatively low yields. In order to increase photosynthetic electron flow towards the production of D-lactate, we have generated several strains of the marine cyanobacterium Synechococcus sp. PCC 7002 (Syn7002) with deletions in genes involved in cyclic or pseudo-cyclic electron flow around photosystem I. Using a variant of the Chlamydomonas reinhardtii D-lactate dehydrogenase (LDH SRT , engineered to efficiently utilize NADPH in vivo), we have shown that deletion of either of the two flavodiiron flv homologs (involved in pseudocyclic electron transport) or the Syn7002 pgr5 homolog (proposed to be a vital part of the cyclic electron transport pathway) is able to increase D-lactate production in Syn7002 strains expressing LDH SRT and the Escherichia coli LldP (lactate permease), especially at low temperature (25°C) and 0.04% (v/v) CO 2 , though at elevated temperatures (38°C) and/or high (1%) CO 2 concentrations, the effect was less obvious. The Dpgr5 background seemed to be particularly beneficial at 25°C and 0.04% (v/v) CO 2 , with a nearly 7-fold increase in D-lactate accumulation in comparison to the wild-type background (≈1000 vs ≈150 mg/L) and decreased side effects in comparison to the flv deletion strains. Overall, our results show that manipulation of photosynthetic electron flow is a viable strategy to increase production of platform chemicals in cyanobacteria under ambient conditions.

Regulatory electron transport pathways of photosynthesis in cyanobacteria and microalgae: Recent advances and biotechnological prospects

“…Loss of Flv1 and/or Flv3 increased D-lactate titers under constant illumination, mild temperature and atm.CO 2 . However, ΔFlv1 and ΔFlv3 strains but not the WT underwent chlorosis during the production(Selão et al, 2020). Importantly, deletion of Pgr5 did not induce chlorosis and led to the highest D-lactate titer observed in the study.…”

“…Application of Synechocystis cells deficient in FDPs and RTOs (ΔFlv2/ΔFlv3/ΔRTOs) increased the power density fivefold (Saar et al, 2018), thus expanding utilization of photosynthetic microbes in emerging technologies. (Selão et al, 2020).…”

“…CO 2 . However, ΔFlv1 and ΔFlv3 strains but not the WT underwent chlorosis during the production (Selão et al, 2020). Importantly, deletion of Pgr5 did not induce chlorosis and led to the highest d ‐lactate titer observed in the study.…”

“…Importantly, deletion of Pgr5 did not induce chlorosis and led to the highest d ‐lactate titer observed in the study. However, elevating either CO 2 level and/or temperature abolished the production advantage of each photosynthetic mutant over the d ‐lactate dehydrogenase‐expressing reference strain (Selão et al, 2020).…”

Gene expression and organization of thylakoid protein complexes in the PSII‐less mutant of Synechocystis sp. PCC 6803

“…Furthermore, protein complexes involved in cyclic electron transfer and carbon concentration mechanisms are also essential components of the thylakoid membrane (Burnap et al, 2013;Lea-Smith et al, 2016;Selão et al, 2020).…”