472567 PHA Production Using Yarrowia Lipolytica and Alternative Feedstocks

Wednesday, November 16, 2016: 8:48 AM
Continental 9 (Hilton San Francisco Union Square)
Michael Spagnuolo, Difeng Gao and Mark A. Blenner, Chemical & Biomolecular Engineering, Clemson University, Clemson, SC

Polyhydroxyalkanoates (PHAs) are naturally-occurring polyesters produced by a number of different bacterial species as a means of carbon and energy storage. PHAs have been suggested as possible replacements and supplements to petroleum-derived plastics due to PHAs’ tunable mechanical properties and high potential for feedstock flexibility. Feedstock flexibility allows production of a desired product even under conditions of environmental or economically-induced scarcity. Natural producers of PHAs are often highly sensitive to the substrates and media on which they are grown. This variability precludes feedstock flexibility. In order to address this issue, we have engineered a far more metabolically-robust organism, the oleaginous yeast Yarrowia lipolytica, to produce PHAs from a number of different feedstocks.

To accomplish this, the PHA-producing enzymes, called PHA Synthases, from the natural PHA producers Pseudomonas aeruginosa and Ralstonia eutropha were incorporated into the yeast. Flux to this enzyme was increased by overexpression of the native acetyl-CoA carboxylase (ACC1) and modification of Multi-Function Enzyme (MFE1). ACC1 facilitates the synthesis of longer carbon chains, which is desirable when building a specific larger product from smaller substrates. The modification to MFE1 prevents this double-acting enzyme from degrading a highly desired PHA precursor. Yarrowia lipolytica’s native catabolic pathways allow it to be an effective metabolizer of fats, sugars, and proteins. Our engineered organisms demonstrated PHA production using pure and waste fats, crude glycerol, and hexose and pentose sugar feedstocks. The final PHA products were also characterized for their structure and composition: necessary information for the tuning of the mechanical properties.

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