Superior thermal stability and fast crystallization behavior of a novel, biodegradable α-methylated bacterial polyester

Abstract

Given their ubiquity in modern society, the development of biodegradable and renewably sourced plastics is essential for the creation of an environmentally sustainable society. One of the drawbacks for currently available biodegradable plastics such as poly(l-lactic acid) (PLLA) and polyhydroxyalkanoates (PHAs) is that it is difficult to simultaneously achieve mechanical flexibility and certain crystallization behavior in these materials, which limits their use as replacements for established petroleum-based plastics such as isotactic polypropylene (iPP). Here, we report the synthesis and characterization of a new biodegradable plastic, poly(3-hydroxy-2-methylbutyrate) [P(3H2MB)], which is a member of the bacterial PHA family whose members include an α-methylated monomer unit. Biosynthesis of P(3H2MB) was achieved using recombinant Escherichiacoli expressing an engineered pathway. Biosynthesized P(3H2MB) exhibited the highest melting temperature (197 °C) among the biosynthesized PHAs and improved thermal resistance. It also exhibited improved crystallization behavior and mechanical flexibility nearly equal to those of iPP. The primary nucleation rate of P(3H2MB) was faster than that of P(3HB), and the spherulite morphology of P(3H2MB) was much finer than that of P(3HB). This crystal morphology may result in more rapid crystallization behavior, increased transparency, and enhanced mechanical properties. The superior physical properties of P(3H2MB) have the potential to open new avenues for the production of high-performance biodegradable plastics for replacing petroleum-based bulk commodity plastics.

Exploring Class I polyhydroxyalkanoate synthases with broad substrate specificity for polymerization of structurally diverse monomer units

“…As PHA production is associated with cell growth ( Sudesh et al, 2000 ), it is imperative to eliminate or reduce the risk of toxicity induced by such precursors. Therefore, the precursors were introduced into the culture medium after 4 h, once substantial cell growth was achieved, mainly for better tolerance ( Furutate et al, 2021 ). Meanwhile, a high glucose concentration can cause catabolic repression of phaJ and pct genes induced by IPTG ( Furutate et al, 2021 ); thus, the glucose concentration was maintained at a minimum to promote cell growth only.…”

“…E. coli LSBJ, a fadB fadJ double-deletion strain of E. coli LS5218 [ fadR601 , atoC (Con)] ( Tappel et al, 2012a ), was used as the host strain for PHA biosynthesis. This strain is an ideal host for non-native PHA production because of its ability to take up a wide variety of substrates to be incorporated into PHA homo- and copolymers, and bench-level scale-up methodologies available for overall production ( Tappel et al, 2012b ; Levine et al, 2016 ; Pinto et al, 2016 ; Fadzil et al, 2018 ; Furutate et al, 2021 ; Scheel et al, 2021 ). A broad-host-range plasmid pBBR1MCS-2 ( Kovach et al, 1995 ) harboring the genes encoding the PhaCs to be evaluated, the lac promoter region, the ( R )-specific enoyl-CoA hydratase gene from A. caviae ( phaJ Ac ), the 3-ketothiolase gene ( phaA ) from Ralstonia eutropha H16, and the acetoacetyl-CoA reductase gene ( phaB ) from R. eutropha H16, termed pBBR1-phaCsAB Re J Ac , was used for the expression of PhaCs ( Supplementary Figure S1 ).…”

“…Inoculations for PHA production were started with 5 mL of seed culture added to 500 mL shake flasks containing 95 mL of modified M9 medium ( Furutate et al, 2021 ) (final volume:100 mL and 5% inoculum). The modified M9 medium comprised of 17.1 g/L Na 2 HPO 4 ·12H 2 O, 3 g/L KH 2 PO 4 , 0.5 g/L NaCl, 2 mL of 1 M MgSO 4 ·7H 2 O, 0.1 mL of 1 M CaCl 2 , and 2.5 g/L Bacto-yeast extract.…”

“…The number of PhaC enzymes with broad substrate specificity is scarce; thus, the new PhaC enzymes reported in this study are highly intriguing for future studies. Furthermore, PHAs containing α-carbon methylated units are potentially attractive biobased materials (Füchtenbusch et al, 1998;Furutate et al, 2021); thus, PhaCs with the ability to polymerize 3H2MB and 3HPi are of great interest. PhaC Ps demonstrated superior performance in the Frontiers in Bioengineering and Biotechnology frontiersin.org polymerization of 3HPi to PhaC Ac .…”

“…The poor material properties of P(3HB) ( Lehrle and Williams, 1994 ) such as its high crystallinity and narrow processing temperature window have greatly hampered the entry of this polymer into the commercial world. Fortunately, 3HB-based copolymers ( Tsuge et al, 2005 ; Mizuno et al, 2010 ; Mierzati et al, 2020 ; Furutate et al, 2021 ) have been proven to overcome the material property limitations of P(3HB) to a certain extent, and have been used as a remedy for problems related to plastics ( Sivashankari and Tsuge, 2021 ).…”