Precision Engineering of the Co-immobilization of Enzymes for Cascade Biocatalysis.

The design and orderly layered co-immobilization of multiple enzymes on resin particles remain challenging. In this study, the SpyTag/SpyCatcher binding pair was fused to the N-terminus of an alcohol dehydrogenase (ADH) and an aldo-keto reductase (AKR), respectively. A non-canonical amino acid (ncAA), p-azido-L-phenylalanine (p-AzF), as the anchor for covalent bonding enzymes, was genetically inserted into preselected sites in the AKR and ADH. Employing the two bioorthogonal counterparts of SpyTag/SpyCatcher and azide-alkyne cycloaddition for the immobilization of AKR and ADH enabled sequential dual-enzyme coating on porous microspheres. The ordered dual-enzyme reactor was subsequently used to synthesize (S)-1-(2-chlorophenyl)ethanol asymmetrically from the corresponding prochiral ketone, enabling the in situ regeneration of NADPH. The reactor exhibited a high catalytic conversion of 74 % and good reproducibility, retaining 80 % of its initial activity after six cycles. The product had 99.9 % ee, which that was maintained in each cycle. Additionally, the double-layer immobilization method significantly increased the enzyme loading capacity, which was approximately 1.7 times greater than that of traditional single-layer immobilization. More importantly, it simultaneously enabled both the purification and immobilization of multiple enzymes on carriers, thus providing a convenient approach to facilitate cascade biocatalysis.

Succinic anhydride-based chemical modification making laccase@Cu3(PO4)2 hybrid nanoflowers robust in removing bisphenol A in wastewater.

To prepare a robust biocatalyst and enhance the removal of bisphenol A in wastewater, succinic anhydride was reacted with laccase to obtain succinic anhydride-modified laccase (SA-laccase) and then co-crystallized with Cu3(PO4)2 to form SA-laccase@Cu3(PO4)2 hybrid nanoflowers (hNFs). The activity of SA-laccase@Cu3(PO4)2 reached 5.27 U/mg, 1.86-, 2.88- and 2.15-fold those of bare laccase@Cu3(PO4)2, laccase@Ca3(PO4)2 and laccase@epoxy resin, respectively. Compared with free laccase, the obtained hNFs present enhanced activity and tolerance to pH and high temperature in the removal of BPA. Under the optimum conditions of pH 6.0 and 35 °C, BPA removal reached 93.2% using SA-laccase@Cu3(PO4)2 hNFs, which was 1.21-fold of that using free laccase. In addition, the obtained SA-laccase@Cu3(PO4)2 hNFs retained nearly 90% of their initial catalytic activity for BPA removal after 8 consecutive batch cycles. This efficient method for preparing immobilized laccase can also be further developed and improved to acquire green biocatalysts for removing persistent organic pollutants in wastewater.

A double-blind, clinical evaluation of facial augmentation treatments: a comparison of PRI 1, PRI 2, Zyplast and Perlane.

BACKGROUND: Facial wrinkles are caused by weakening of the sub-dermal collagen support layer. Dermal fillers can be used to treat wrinkles, and this double-blind, randomised, single-centre study compared four fillers: PRI 1, PRI 2 (both porcine collagen), Zyplast (purified bovine collagen) and Perlane (cross-linked hyaluronic acid gel). METHODS: 79 females (aged 25-55 years) with wrinkles in the upper lip line border were randomised to PRI 1 (19 patients), PRI 2 (19 patients), Perlane (23 patients), Zyplast (18 patients). Patients were assessed at 1 week and 1, 3, 6, 9 and 12 months using 2D images and by mathematically derived facial volume changes using 3D stereophotogrammetry. RESULTS: All treatments produced larger, less wrinkled, more prominent lips. PRI 1, PRI 2 and Zyplast showed similar lip volume gains, with Perlane showing the greatest upper lip volume increase. All treatments were comparable for rates of decrease in upper lip volume post-treatment, however, Perlane maintained higher lip volume gains at each time point. Investigators indicated PRI 1 was significantly easier to deliver than Zyplast. Patient satisfaction scores were similar, though there was a trend towards greater dissatisfaction for PRI 1 and PRI 2 at month 9 (p=0.052). Treatment was well-tolerated, with 'cold sore' being the most common adverse event. CONCLUSIONS: Results showed that PRI 1 and PRI 2 were comparable to Perlane and Zyplast as dermal fillers. Further rigorous studies are required to establish the performance of dermal fillers and patient acceptability. We propose the utilisation of stereophotogrammetry for assessment of volume changes.

A new three-dimensional method of assessing facial volumetric changes after orthognathic treatment.

OBJECTIVE: To validate a new method of facial volumetric assessment that is dependent on the use of stereophotogrammetric models and a software-based Facial Analysis Tool. DESIGN: The method was validated in vitro with three-dimensional (3D) models of a lifelike plastic female dummy head and in vivo with a male-subject head. METHODS: Thirty facial silicone explants were added in the nasal and perioral regions of each head, and their volumes were obtained by three different algorithms. These were compared with the actual values obtained by a "water displacement" method. RESULTS: The least mean error was found with the "tetrahedron formation" method followed by the "projection" method and the "back-plane construction" method. The error with the tetrahedron formation method was 0.071 cm(3) (95% confidence interval [CI]: -0.074 to 0.2161 cm3) with the in vitro models and 0.314 cm3 (95% CI: -0.080 to 0.708 cm3) with the in vivo models. The increased volumetric assessment error observed in vivo was attributed to the registration procedure and possible changes in facial expression. CONCLUSIONS: These results encourage the use of this method in the 3D assessment of orthognathic surgical outcome, provided a standardized facial expression is used for image acquisition.