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For a sustainable economy, biodegradable biopolymers polyhydroxyalkanoates (PHA) are desirable substitutes to petroleum-based plastics that contaminate our environment. Medium-chain-length (MCL) PHA bioplastics are particularly interesting due to their thermoplastic properties. To hamper the high cost associated to PHA production, the use of bacterial mixed cultures cultivated in open systems and using cheap resources is a promising strategy. Here, we studied the operating conditions favouring direct MCL accumulation by activated sludge, using oleic acid as a model substrate and phosphorus limitation in fed-batch bioreactors. Our results confirm the presence of PHA-accumulating organisms (PHAAO) in activated sludge able to accumulate MCL from oleic acid. A positive correlation between phosphorus (P) limitation and PHA accumulation was demonstrated, allowing up to 26% PHA/total biomass accumulation, and highlighted its negative impact on the MCL/PHA fraction in the polymer. Diversity analysis through 16S rRNA amplicon sequencing revealed a differential selection of PHAAO according to the P-limitation level. A differential behaviour for the orders Pseudomonadales and Burkholderiales at increasing P-limitation levels was revealed, with a higher abundance of the latter at high levels of P-limitation. The PHA accumulation observed in activated sludge open new perspectives for MCL-PHA production system based on P-limitation strategy applied to mixed microbial communities. Key points: • Direct accumulation of MCL-PHA in activated sludge was demonstrated. • MCL-PHA content is negatively correlated with P-limitation. • Burkholderiales members discriminate the highest P-limitation levels. [ABSTRACT FROM AUTHOR]

Polyhydroxyalkanoates (PHAs) are biopolymers synthesized by a wide range of bacteria, which serve as a promising candidate in replacing some conventional petrochemical-based plastics. PHA synthase (PhaC) is the key enzyme in the polymerization of PHA, and the crystal structures were successfully determined using the catalytic domain of PhaC from Cupriavidus necator (PhaCCn-CAT) and Chromobacterium sp. USM2 (PhaCCs-CAT). Here, we review the beneficial mutations discovered in PhaCs from a structural perspective. The structural comparison of the residues involved in beneficial mutation reveals that the residues are near to the catalytic triad, but not inside the catalytic pocket. For instance, Ala510 of PhaCCn is near catalytic His508 and may be involved in the open-close regulation, which presumably play an important role in substrate specificity and activity. In the class II PhaC1 from Pseudomonas sp. 61-3 (PhaC1Ps), Ser325 stabilizes the catalytic cysteine through hydrogen bonding. Another residue, Gln508 of PhaC1Ps is located in a conserved hydrophobic pocket which is next to the catalytic Asp and His. A class I, II-conserved Phe420 of PhaCCn is one of the residues involved in dimerization and its mutation to serine greatly reduced the lag phase. The current structural analysis shows that the Phe362 and Phe518 of PhaC from Aeromonas caviae (PhaCAc) are assisting the dimer formation and maintaining the integrity of the core beta-sheet, respectively. The structure-function relationship of PhaCs discussed in this review will serve as valuable reference for future protein engineering works to enhance the performance of PhaCs and to produce novel biopolymers. [ABSTRACT FROM AUTHOR]

During the last decades, biopolymers experienced a renaissance. The increasing limitation of fossil resources in combination with a public demand for environmental-friendly and sustainable processes has led to the formation of a market for biobased plastics. Especially non-biodegradable bioplastics are very interesting materials, as they combine the benefits of reduced carbon footprint during production and increased resource efficiency with the persistence to microbial degradation. Consequently, persistent biomass-derived plastic materials are highly promising to substitute conventional fossil-based plastics in applications, which require durability and longevity. Non-biodegradable bioplastics derived from renewable resources represent 57% of all bioplastics with partially biobased polyethylene terephthalate currently leading the market, followed by biobased polyamides and fully biomass-derived polyethylene. An exceptional biopolymer with thermoplastic properties was discovered only two decades ago, when—for the first time—polythioesters were synthesized by microbial fermentation. Though synthesized by bacteria, it turned out that polythioesters are non-biodegradable by microorganisms in contrast to all other biopolymers and thus, represent a novel non-biodegradable bioplastic material. This review gives an overview about the recent development and progress regarding bioplastics with special focus on persistent bioplastics. We describe the generation of the respective monomers from biomass-derived substrates and summarize the current status of production, which range from the laboratory-scale up to large-scale industrial processes. [ABSTRACT FROM AUTHOR]

Poly-3-hydroxyalkanoates [P(3HA)s] are biologically produced polyesters that have attracted much attention as biodegradable polymers that can be produced from biorenewable resources. These polymers have many attractive properties for use as bulk commodity plastics, fishing lines, and medical uses that are dependent on the repeating unit structures. Despite the readily apparent benefits of using P(3HA)s as replacements for petrochemical-derived plastics, the use and distribution of P(3HA)s have been limited by their cost of production. This problem is currently being addressed by the engineering of enzymes involved in the production of P(3HA)s. Polyhydroxyalkanoate (PHA) synthase (PhaC) enzymes, which catalyze the polymerization of 3-hydroxyacyl-CoA monomers to P(3HA)s, were subjected to various forms of protein engineering to improve the enzyme activity or substrate specificity. This review covers the recent history of PHA synthase engineering and also summarizes studies that have utilized engineered PHA synthases. [ABSTRACT FROM AUTHOR]

Poly 3-hydroxybutyric acid (PHB) and related copolymers can be advantageously produced when integrated into a sugarcane mill. In this favorable scenario, the energy necessary for the production process is provided by biomass. Carbon dioxide emissions to the environment are photosynthetically assimilated by the sugarcane crop and wastes are recycled to the cane fields. The polymer can be produced at low cost considering the availability of a low-price carbon source and energy. [ABSTRACT FROM AUTHOR]