Page 87 - Biodegradable Polyesters

3.8 Application Opportunities of Bacterial Polyhydroxyalkanoates 65

to the medium to synthesize P(LA-co-3HV). As a result, P(LA-co-3HB-co-3HV)
terpolymer with 96 mol% LA was obtained. These findings demonstrated that
the very weak supply of 3HB-CoA and/or 3HV-CoA from valerate achieved
the production of the LA-enriched polymer. Thus, only 4 mol% of secondary
monomer units was required for the incorporation of the LA units. Worthy of
note is that the incapability of PLA homopolymer synthesis is not due to the
inability to avoid the presence of 3HB-CoA because no polymer was obtained
without 3HB-CoA-supplying enzymes as mentioned above.

3.8.7
Applications of PHA Inclusions as Functionalized Biobeads

3.8.7.1 Bioseparations
PHB biobeads displaying the ZZ domain of Protein A from Staphylococcus aureus
as the result of N-terminal fusion to PhaC were found to be suitable to purify
IgG from serum samples and culture supernatant with high binding capacity and
purification power [20, 68]. Other binding domains were successfully displayed
such as scFv (single-chain variable fragment) or streptavidin and enabled applica-
tion of the respective beads as affinity purification resin [69, 70].
Recently, PHB beads for endotoxin removal were developed by fusing the
human lipopolysaccharide binding protein (hLBP) to PhaP immobilized in vitro
on PHB particles via the natural hydrophobic interaction between PhaP and PHB
[71].

3.8.7.2 Drug Delivery
As mentioned above, PLA and poly(lactic-co-glycolic acid) have been used for
drug delivery for many years and PHAs have only recently attracted interest for
use in similar applications, particularly as nano-size materials. For example, PHA
nanoparticles were loaded with rhodamine B isothiocyanate (RBITC) in vitro and
then targeted to macrophages. RBITC was released over period of 20 days, while
macrophage viability was maintained [72]. PHA nanoparticles could be loaded
with the phosphoinositide-3-kinases (PI3Ks) inhibitor and were able to inhibit
proliferation of cancer cell lines, suggesting applicability of PHA particles in can-
cer therapy [73].

3.8.7.3 Protein Purification
PHA beads have been demonstrated for their utility to purify proteins imple-
menting two very different approaches. One approach uses the production of
recombinant target proteins as fusion proteins to be attached to the surface of
PHA inclusions during formation and the second approach considered PHA
beads which display a binding domain with binding affinity to the target.
PHA bead-based fusion protein purification strategies have utilized fusions of
the target protein to proteins naturally associated with PHA inclusions such as
phasins, regulatory, or synthase proteins. Banki et al. [74] used a phasin–intein
target protein fusion to anchor the fusion protein to the PHA granule as it forms