Ethylene production with engineered Synechocystis sp PCC 6803 strains

Microbial Cell Factories

Research

Klaas J. Hellingwerf¹  S. Andreas Angermayr²  Vinod Puthan Veetil³ 

[1] Molecular Microbial Physiology Group, Swammerdam Institute for Life Sciences, University of Amsterdam, Science Park 904, 1098 XH, Amsterdam, The Netherlands;Molecular Microbial Physiology Group, Swammerdam Institute for Life Sciences, University of Amsterdam, Science Park 904, 1098 XH, Amsterdam, The Netherlands;Institute of Science and Technology (IST) Austria, Klosterneuburg, Austria;Molecular Microbial Physiology Group, Swammerdam Institute for Life Sciences, University of Amsterdam, Science Park 904, 1098 XH, Amsterdam, The Netherlands;Reliance Technology Group, Reliance Industries Ltd., Navi Mumbai, India;

关键词: Ethylene;    Cyanobacteria;    Sustainable;    Synechocystis;    Glycogen;    Oxoglutarate;    Arginine;   

DOI  :  10.1186/s12934-017-0645-5

 received in 2016-10-23, accepted in 2017-02-09,  发布年份 2017

来源: Springer

【 摘 要 】

BackgroundMetabolic engineering and synthetic biology of cyanobacteria offer a promising sustainable alternative approach for fossil-based ethylene production, by using sunlight via oxygenic photosynthesis, to convert carbon dioxide directly into ethylene. Towards this, both well-studied cyanobacteria, i.e., Synechocystis sp PCC 6803 and Synechococcus elongatus PCC 7942, have been engineered to produce ethylene by introducing the ethylene-forming enzyme (Efe) from Pseudomonas syringae pv. phaseolicola PK2 (the Kudzu strain), which catalyzes the conversion of the ubiquitous tricarboxylic acid cycle intermediate 2-oxoglutarate into ethylene.ResultsThis study focuses on Synechocystis sp PCC 6803 and shows stable ethylene production through the integration of a codon-optimized version of the efe gene under control of the Ptrc promoter and the core Shine–Dalgarno sequence (5′-AGGAGG-3′) as the ribosome-binding site (RBS), at the slr0168 neutral site. We have increased ethylene production twofold by RBS screening and further investigated improving ethylene production from a single gene copy of efe, using multiple tandem promoters and by putting our best construct on an RSF1010-based broad-host-self-replicating plasmid, which has a higher copy number than the genome. Moreover, to raise the intracellular amounts of the key Efe substrate, 2-oxoglutarate, from which ethylene is formed, we constructed a glycogen-synthesis knockout mutant (ΔglgC) and introduced the ethylene biosynthetic pathway in it. Under nitrogen limiting conditions, the glycogen knockout strain has increased intracellular 2-oxoglutarate levels; however, surprisingly, ethylene production was lower in this strain than in the wild-type background.ConclusionMaking use of different RBS sequences,production of ethylene ranging over a 20-fold difference has been achieved. However, a further increase of production through multiple tandem promoters and a broad-host plasmid was not achieved speculating that the transcription strength and the gene copy number are not the limiting factors in our system.

【 授权许可】

CC BY   
© The Author(s) 2017