Metabolic engineering of rapidly growing <scp> <i>Synechococcus elongatus</i> </scp> strains for phototrophic production of alkanes

Alkanes are high-energy hydrocarbons that are foreseen as next generation biofuels. Cyanobacteria are known to naturally synthesize C15-C19 alkanes; however, the titers are too low to make this a commercially viable process. Therefore, to leverage these photosynthetic platforms for improved alkane production, here we engineered three novel isolates of Synechococcus elongatus PCC 11801, PCC 11802, and IITB6. The two gene AAR-ADO alkane biosynthesis pathway was constructed by cloning the genes for acyl-ACP reductase (aar) and aldehyde deformylating oxygenase (ado) from S. elongatus PCC 7942 under the regulation of P_rbcL promoter from PCC 7942 and native promoters from PCC 11801 such as P_cpcB300, P_psbAI, and P_psbAIII. The genes were separately cloned under two different promoters, creating a library of the engineered strains. The results indicated that the engineered strains of novel S. elongatus isolates produced significantly higher amounts of alkanes than the model strain PCC 7942. The highest alkane yield achieved was 4.1 mg/gDCW in BG-11, while the highest titer was 31.5 mg/L in 5X BG-11, with an engineered IITB6 strain (P_cpcB300:aar:T_rrnB::P_rbcL:ado:T_Lac). Overall, the study highlights the potential of newly isolated S. elongatus strains as efficient alkane production platforms.