Cellulose Nanocrystals (CNCs) and Its Modified Form from Durian Rind as Dexamethasone Carrier.

In this study, CNCs were extracted from durian rind. Modification to CNCs with saponin was conducted at 50 °C for one h. CNCs and CNCs-saponin were employed as dexamethasone carriers. Modification to CNCs using saponin did not change the relative crystallinity of CNCs. CNCs' molecular structure and surface chemistry did not alter significantly after modification. Both nanoparticles have surface charges independently of pH. Dexamethasone-released kinetics were studied at two different pH (7.4 and 5.8). Higuchi, Ritger-Peppas, first-order kinetic and sigmoidal equations were used to represent the released kinetic data. The sigmoidal equation was found to be superior to other models. The CNCs and CNCs-saponin showed burst release at 30 min. The study indicated that cell viability decreased by 30% after modification with saponin.

Class I hydrophobin fusion with cellulose binding domain for its soluble expression and facile purification.

Hydrophobins, highly surface-active proteins, have the ability to reverse surface hydrophobicity through self-assembly at the hydrophilic-hydrophobic interfaces. Their unique structure and interfacial activity lead hydrophobins to have potential applications on surface functional modifications. However, class I hydrophobins are prone to self-assemble into highly insoluble amyloid-like rodlets structure. Recombinant hydrophobins could be produced by Escherichia coli but generally as an insoluble inclusion body. To overcome this insoluble expression limitation, cellulose-binding domain (CBD) from Clostridium thermocellum was fused to the N-terminal of class I hydrophobin HGFI to enhance its soluble expression in E. coli. Approximately, 94% of expressed CBD fused HGFI (CBD-HGFI) was found as soluble protein. The fused CBD could also bind specifically onto bacterial cellulose (BC) nanofibrils produced by Komagataeibacter xylinus to facilitate rapid isolation and purification of HGFI from crude extract. Lysostaphin (Lst), known as GlyGly endopeptidase could successfully cleave the flexible linker (GGGGS)2 between CBD and HGFI to recover HGFI from BC-bound CBD-HGFI. CBD-HGFI purified by immobilized metal-chelated affinity chromatography (IMAC) and Lst cleaved BC-CBD-HGFI still retained interfacial activity of hydrophobin and its effect on accelerating PETase hydrolysis against poly(ethylene terephthalate) (PET) fiber.

Class I hydrophobins pretreatment stimulates PETase for monomers recycling of waste PETs.

Poly(ethylene terephthalate) hydrolase (PETase) from Ideonella sakaiensis 201-F6 was expressed and purified from Escherichia coli to hydrolyze poly(ethylene terephthalate) (PET) fibers waste for its monomers recycling. Hydrolysis carried out at pH 8 and 30 °C was found to be the optimal condition based on measured monomer mono(2-hydroxyethyl) terephthalate (MHET) and terephthalic acid (TPA) concentrations after 24 h reaction. The intermediate product bis(2-hydroxyethyl) terephthalate (BHET) was a good substrate for PETase because BHET released from PET hydrolysis was efficiently converted into MHET. Only a trace amount of MHET could be further hydrolyzed to TPA. Class I hydrophobins RolA from Aspergillus oryzae and HGFI from Grifola frondosa were expressed and purified from E. coli to pretreat PET surface for accelerating PETase hydrolysis against PET. The weight loss of hydrolyzed PET increased from approximately 18% to 34% after hydrophobins pretreatment. The releases of TPA and MHET from HGFI-pretreated PET were enhanced 48% and 62%, respectively. The selectivity (TPA/MHET ratio) of the hydrolysis reaction was approximately 0.5.

Fungal Hydrophobin RolA Enhanced PETase Hydrolysis of Polyethylene Terephthalate.

Polyethylene terephthalate (PET) becomes one of the most well-known polyesters and is widely used as packaging material. Recently, polyethylene terephthalate hydrolase (PETase) has emerged as a potential biocatalyst demonstrating the ability to degrade polyethylene terephthalate (PET). We showed that the rate of PETase hydrolysis could be significantly increased in the presence of hydrophobin RolA. Hydrophobins represent a class of small fungal protein that has a high surface-active substance and can spontaneously self-assemble at hydrophilic-hydrophobic interfaces. In this work, a class I hydrophobin named RolA was extracted from the mycelium pellet collected from a fermentation culture of Aspergillus oryzae. The SDS-PAGE analysis of the isolated RolA showed the presence of 11 kDa polypeptide. Recombinant PETase from Ideonella sakaiensis was also successfully expressed in Escherichia coli as a soluble protein with molecular weight approximately 30 kDa. The hydrophobin RolA could enhance the PET hydrolysis in the presence of the recombinant PETase. The hydrolysis of PET bottle by RolA-PETase achieved the highest weight loss of 26% in 4 days. It is speculated that the wetting effect of RolA acts on PET surface converts PET to become hydrophilic that leads PETase easier to contact and attack the surface. Graphical Abstract.