Investigation of Isoprene Synthase Protein Expression in Synechococcus sp. PCC 7002 by Liquid Chromatography Tandem-Mass Spectrometry (LC-MS/MS)

Biofuels are hydrocarbons and like fossil fuels create water and CO₂ when combusted. Carbon dioxide is a greenhouse gas that traps heat in the atmosphere contributing to global warming. However, when photosynthesis is used to produce biofuels, the carbon is taken from CO₂ that was already in the atmosphere. Isoprene (C₅H₈) is a small volatile hydrocarbon, used as a feedstock for synthetic rubber, and can be used for production of liquid fuels. For isoprene synthesis the methyl-erythritol-4-phosphate (MEP) pathway is needed. Formation of isoprene is due to the presence of an isoprene synthase (IspS) gene. Cyanobacteria are not gifted with the IspS gene, yet they do express the MEP pathway. Synechococcus sp. PCC 7002 is a marine cyanobacterium; that is fast growing and naturally transformable, making it ideal for isoprene synthesis if the IspS gene can be inserted and functionally expressed. The objective of this project is to detect and quantify the levels of the IspS enzyme in Synechococcus sp. PCC 7002 mutants that have been genetically engineered to express introduced IspS genes. In the data obtained it has not yet been shown that IspS enzymes have been synthesized and that these catalyze the production of isoprene. The intent of my project is to determine whether the introduced IpsS genes have been expressed at the protein level by Liquid Chromatography Tandem-Mass Spectrometry (LC-MS/MS) and whether isoprene can be detected. Future work can focus on optimization of isoprene production in transformed Synechococcus sp. PCC 7002 for commercially viable isoprene production projects.