Disruption of β-oxidation pathway in Pseudomonas putida KT2442 to produce new functionalized PHAs with thioester groups

Abstract

This work describes the generation of novel PHAs (named PHACOS) with a new monomer composition containing thioester groups in the side chain, which confers new properties and made them suitable for chemical modifications after their biosynthesis. We have analyzed the PHACOS production abilities of the wild-type strain Pseudomonas putida KT2442 vs. its derived strain P. putida KT42FadB, mutated in the fadB gene from the central metabolic β-oxidation pathway involved in the synthesis of medium-chain-length PHA (mcl-PHA). Different fermentation strategies based on one- or two-stage cultures have been tested resulting in PHACOS with different monomer composition. Using decanoic acid as inducer of the growth and polymer synthesis and 6-acetylthiohexanoic acid as PHA precursor in a two-stage strategy, the maximum yield was obtained by culturing the strain KT42FadB. Nuclear magnetic resonance and gas chromatography coupled to mass spectrometry showed that polymers obtained from the wild-type and KT42FadB strains, included 6-acetylthio-3-hydroxyhexanoic acid (OH-6ATH) and the shorter derivative 4-acetylthio-3-hydroxybutanoic acid (OH-4ATB) in their composition, although in different ratios. While the polymer obtained from KT42FadB strain contained mainly OH-6ATH monomer units, mcl-PHA produced by the wild-type strain contained OH-6ATH and OH-4ATB. Furthermore, polyesters showed differences in the OH-alkyl derivates moiety. The strain KT42FadB overproduced PHACOS when compared to the production rate of the control strain in one- and two-stage cultures. Thermal properties obtained by differential scanning calorimetry indicated that both polymers have different glass transition temperatures related to their composition.

Novel extracellular medium-chain-length polyhydroxyalkanoate depolymerase from Streptomyces exfoliatus K10 DSMZ 41693: a promising biocatalyst for the efficient degradation of natural and functionalized mcl-PHAs

“…(Valencia, Spain), and PHACOS [P(OH-Alk-co-OH-6ATH-OH4ATB)] was obtained as previously described (Escapa et al 2011). Chromatography media were obtained from GE-Healthcare (Uppsala, Sweden).…”

“…In this sense, introduction of thioester linkages in the polymer backbone has led to different sclPHAs, referred to as polythioesters (PTEs), with novel physical properties (solubility, crystallinity, and thermal stability) (Lütke-Eversloh et al 2001Lütke-Eversloh and Steinbüchel 2003;Kawada et al 2003). Furthermore, it has been recently described a new second-generation polymer containing thioester groups in the side chain (PHACOS) showing tunable thermal and antimicrobial properties (Escapa et al 2011;Dinjaski et al 2014). Since PHAs are composed of enantiopure hydroxyalkanoic acids (RHAs), they are a source of valuable synthons for the chemical and pharmaceutical industries (Ren et al 2010;Chen 2011;Gao et al 2011).…”

Start a Research on Biopolymer Polyhydroxyalkanoate (PHA): A Review

“…Depending on the total number of carbon atoms within a PHA monomer, PHA can be classified as either short-chain length PHA (scl-PHA; 3 to 5 carbon atoms), medium-chain length PHA (mcl-PHA; 6 to 14 carbon atoms), or long-chain length PHA (lcl-PHA; 15 or more carbon atoms) [3]. About 150 different PHA monomers have been identified and this number continues to increase with the introduction of new types of PHA through the chemical or physical modification of naturally-occurring PHA [5], or through the creation of genetically-modified organisms (GMOs) to produce PHA with specialized functional groups [6]. These features gave rise to diverse PHA properties which can be tailored for various applications ranging from biodegradable packaging materials to medical products.…”

Application of Poly(hydroxyalkanoate) In Food Packaging: Improvements by Nanotechnology

“…Among PHAs, copolymers of poly(3-hydroxybutyrate-co-3-hydroxyhexanoate), PHBHx, with low 3HHx residue are identified as one of the most useful members (if compared to PHB) due to its suitable mechanical properties for application as flexible films 85 . An investigation of Escapa et al showed production of poly(hydroxy-6-acetylthiohexanoateco-4-acetylthiobutanoate) produced by P. putida KT2442 on a mixture of 6-acetylthiohexanoic acid 86 .…”