Enzymatic production of clickable and PEGylated recombinant polyhydroxyalkanoates

Polyhydroxyalkanoates (PHAs) are microbial polyesters attracting great interest while further functionalisation could widen their applicability. Various E coli based production systems using the PHA biosynthetic operon from Bacillus cereus 6E/2, were designed to enhance the accumulation of PHAs with high mcl moieties. Media optimization and system engineering were applied, yielding to the production of up to 260 mg/L of PHAs. Polymers characterization revealed a low grade of crystallinity and remarkable hydrophobic features. For further functionalization, a novel enzyme based strategy was developed. Lipase B from Candida antarctica (CaLB) was used to catalyze the terminal coupling of PHA with: i) dimethyl itaconate (DMI) in order to introduce reactive side chain vinyl moieties for easy coupling of functional molecules and/or ii) biocompatible polyethylene glycol (PEG) to tune polymer hydrophilicity. The functionalized DMI-PHA, PEG-PHA and PEG-DMI-PHA polymers obtained and characterised by NMR, GPC, FT-IR, and WCA in this study open new perspectives for the use of PHAs as biodegradable and biocompatible materials of choice for biomedical applications.