| Vascular networks are required to support the formation and function of three-dimensional tissues. Biodegradable scaffolds are being considered in order to promote vascularization where natural regeneration of lost or destroyed vascular networks fails. Particularly; composite materials are expected to fulfill the complex demands of a patient's body to support wound healing. Microbial biopolyesters are being regarded as such second and third generation biomaterials. Methylobacterium extorquens is one of several microorganisms that should be considered for the production of advanced polyhydroxyalkanoates (PHAs). M. extorquens displays a distinct advantage in that it is able to utilize methanol as an inexpensive substrate for growth and biopolyester production.;This project contributed to current research on polyhydroxyalkanoates at different levels. The terminal double bonds of the functionalized scl/mcl-PHAs are amenable to chemical modifications and could be transformed into reactive functional groups for covalently linking other biomacromolecules. It is anticipated that these biopolyesters will be utilized as tissue engineering materials in the future, due to their functionality and thermo-mechanical properties.;Keywords: biopolyesters, functionalized polyhydroxyalkanoates, Methylobacterium extorquens, genetic modification, fermentation in pilot-scale operators, material characterization, thermo-mechanical properties, tissue engineering;The design of functionalized PHAs, which would be made of both saturated short-chain-length (scl, C ≤ 5) and unsaturated medium-chain-length (mcl, 6 ≤ C ≤ 14) monomeric units, aimed at combining desirable material properties of inert scl/mcl-PHAs with those of functionalized mcl-PHAs. By independently inserting the phaC1 or the phaC2 gene from Pseudomonas fluorescens GK13, recombinant M. extorquens strains were obtained which were capable of producing PHAs containing C-C double bonds. A fermentation process was developed to obtain gram quantities of biopolyesters employing the recombinant M. extorquens ATCC 55366 strain which harbored the phaC2 gene of P. fluorescens GK13, the better one of the two strains at incorporating unsaturated monomeric units. The PHAs produced were found in a blend of scl-PHAs and functionalized scl/mcl-PHAs (4 ≤ C ≤ 6), which were the products of the native and of the recombinant PHA synthase, respectively. Thermo-mechanical analysis confirmed that the functionalized scl/mcl-PHAs exhibited the desirable material properties expected. |
