Single cell assessment of yeast metabolic engineering for enhanced lipid production using Raman and AFM-IR imaging

• DOI: 10.1186/s13068-018-1108-x
• Publication Date: 2018-04-19T15:19:53Z
• Authors: Kamila Kochan; Huadong Peng; Bayden R. Wood; Victoria S. Haritos
• Keywords: Metabolic Engineering; Organelle; Lipid droplet

Biodiesel is a valuable renewable fuel made from derivatized fatty acids produced in plants, animals, and oleaginous microbes. Of the latter, yeasts are of special interest due to their wide use biotechnology, ability synthesize store large amounts triacylglycerols while utilizing non-food carbon sources. While yeast efficiently produce lipids, genetic modification indeed, lipid pathway metabolic engineering, usually required for cost-effective production. Traditionally, gas chromatography (GC) used measure acid production track success engineering strategy microbial culture; here we have employed vibrational spectroscopy approaches at population single cell level engineered simultaneously investigating metabolite levels subcellular structures. Firstly, strong correlation (r2 > 0.99) was established between Fourier transform infrared (FTIR) intact cells GC analysis methyl esters differently strains. Confocal Raman individual carrying modifications enhance synthesis accumulation revealed changes body (LB), storage organelle lipids yeast, with number increasing markedly (up tenfold higher); LB size almost double strain that also expressed stabilizing gene but considerable variation noted cells. clear trend toward reduced unsaturated content more complex engineering. Atomic force microscopy-infrared (AFM-IR) indicated differences constituents strains: most highly had elevated much carbohydrate cytoplasm compared unmodified Vibrational allowed simultaneous measurement variability impact on cellular structures as result different introductions or knockouts, within these inform next steps comprehensive Additionally, spectroscopic measures heterogeneity across cultures under modification, an emerging issue efficient biotechnological