Chemical and biological catalysis for the synthesis and recycling of polyhydroxyalkanoates [electronic resource]

An opportunity exists to utilize both methane and wastewater as a source of chemical intermediates, monomers and polymers. This work describes methods that combine molecular synthesis and catalysis with synthetic biology to generate new sustainable materials. The aims of the work are twofold: 1. to develop strategies for poly(hydroxyalkanoate) (PHA) production by methanotrophic bacteria (Chapters 2-4), and 2. to use PHAs as an essentially renewable resource for the production of new chemicals and materials (Chapters 5-6). To meet these aims, studies in four key project areas will be described: (A) the use of new substrates for PHA production in methanotrophs, (B) a combined chemical--biological approach for medium to short chain-length PHA recycling, (C) 2-alkenoate dimerization chemistry, and (D) the synthesis of new materials from 2-alkenoates. PHAs are sustainable alternatives to non-biodegradable petroleum-based plastics, and can be microbially produced by certain types of either heterotrophic or methanotrophic bacteria. Prior to the commencement of this work, obligate methanotrophs capable of producing PHAs under nutrient-limited conditions were limited to the synthesis of poly(3-hydroxybutyrate) (P3HB). Diversifying the range of PHAs available from methanotrophs would greatly expand the range of PHA applications. Chapter 2 reports the first pure culture evidence of methanotrophic synthesis of poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (P(3HB-co-3HV), often abbreviated to PHBV). When grown with methane as sole substrate, the Type II obligate methanotroph Methylocystis parvus OBBP produces P3HB, but not PHBV, under nutrient-limiting conditions. However, when strain OBBP was incubated with C1 substrates (methane, methanol, or formate) and a propionate or valerate co-substrate, synthesis of PHBV was observed. PHBV production was confirmed by gas chromatography, gel permeation chromatography, differential scanning calorimetry, and nuclear magnetic resonance with [13C1]-valerate. Chapter 3 describes further expansion of the range of PHAs available from obligate methanotrophs: by incorporating ω-hydroxyacid co-substrates under nutrient-limiting conditions, poly(3-hydroxybutyrate-co-4-hydroxybutyrate), poly(3-hydroxybutyrate-co-5-hydroxvalerate-co-3-hydroxyvalerate) and poly(3-hydroxybutyrate-co-6-hydroxcaproate-co-4-hydroxybutyrate) can be formed. For any polymer, end-of-life issues are important considerations. Medium chain-length PHAs are emerging products in the PHA market, and Chapter 4 demonstrates that these PHAs can be selectively depolymerized to their constituent monomers through base-catalyzed pyrolysis. The pyrolysis products can be fed back to the methanotrophic bacteria as a co-substrate with methane to recycle the products into short chain-length PHAs. Labeling studies have shed light on the mechanism of the incorporation of these pyrolysis products in methanotrophs. Chapter 5 considers synthetic chemistry that can be done with the products of PHA pyrolysis, namely 2-alkenoic acids. The simplest 3-alkyl-alk-2-enoic is crotonic acid, produced via P3HB depolymerization, but making chemical intermediates and materials using crotonates is a fundamental challenge in chemistry. Chapter 5 details a mild and rapid dimerization of 2-alkenoates to produce new monomers that can be used in novel step-growth polymerizations. Chapter 6 exploits this dimerization strategy to create novel organogels and polymers, and also details attempts at polymerizing crotonates.

Keywords: Chemistry.
Advisor: Waymouth Robert M. primary advisor. Thesis advisor
Criddle Craig advisor. Thesis advisor
Wender Paul A. advisor. Thesis advisor