Investigation of targeting inhibitors of Aβ aggregation, heme-Aβ peroxidase-like activity and efficient detectors of Aβ aggregation, are of therapeutic value and diagnostics significance for the treatment of Alzheimer's disease (AD). Due to the complex pathogenesis of AD, theranostics treatment with multiple functions are necessary. Herein we constructed the NIR absorption property of gold nanorods (GNRs) loaded with single chain variable fragment ( scFv ) 12B4 and thermophilic acylpeptide hydrolase (APH) ST0779 as a smart theranostic complex (GNRs-APH- scFv , GAS), which possesses both rapid detection of Aβ aggregates and NIR photothermal treatment that effectively disassembles Aβ aggregates and inhibits Aβ-mediated toxicity. Methods : We screened targeting anti-Aβ scFv 12B4 and thermophilic acylpeptide hydrolase as amyloid-degrading enzyme, synthesized GAS gold nanorods complex. The GAS was evalued by Aβ inhibition and disaggregation assays, Aβ detection assays, Aβ mediated toxicity assays in vitro . In vivo, delaying Aβ-induced paralysis in AD model of Caenorhabditis elegans was also tested by GAS. Results : In vitro , GAS has a synergistic effect to inhibit and disassociate Aβ aggregates, in addition to decrease heme-Aβ peroxidase-like activity. In cultured cells, treatment with GAS reduces Aβ-induced cytotoxicity, while also delaying Aβ-mediated paralysis in CL4176 C.elegans model of AD. Furthermore, the photothermal effect of the GAS upon NIR laser irradiation not only helps disassociate the Aβ aggregates but also boosts APH activity to clear Aβ. The GAS, as a targeting detector and inhibitor, allows real-time detection of Aβ aggregates. Conclusion : These results firstly highlight the combination of scFv , APH and nanoparticles to be theranostic AD drugs. Taken together, our strategy provides a new thought into the design of smart compounds for use as efficiently therapeutic and preventive agents against AD. Moreover, our design provides broad prospects of biomedical strategy for further theranostics application in those diseases caused by abnormal protein.

Pyrophosphate (PPi) is a byproduct of over 120 biosynthetic reactions, and an overabundance of PPi can inhibit industrial synthesis. Pyrophosphatases (PPases) can effectively hydrolyze pyrophosphate to remove the inhibitory effect of pyrophosphate. In the present work, a thermophilic alkaline inorganic pyrophosphatase from Thermococcus onnurineus NA1 was studied. The optimum pH and temperature of Ton1914 were 9.0 and 80 °C, respectively, and the half-life was 52 h at 70 °C and 2.5 h at 90 °C. Ton1914 showed excellent thermal stability, and its relative enzyme activity, when incubated in Tris-HCl 9.0 containing 1.6 mM Mg2+ at 90 °C for 5 h, was still 100%, which was much higher than the control, whose relative activity was only 37%. Real-time quantitative PCR (qPCR) results showed that the promotion of Ton1914 on long-chain DNA was more efficient than that on short-chain DNA when the same concentration of templates was supplemented. The yield of long-chain products was increased by 32–41%, while that of short-chain DNA was only improved by 9.5–15%. Ton1914 also increased the yields of UDP-glucose and UDP-galactose enzymatic synthesis from 40.1% to 84.8% and 20.9% to 35.4%, respectively. These findings suggested that Ton1914 has considerable potential for industrial applications.

Enzyme activation is a powerful means of achieving biotransformation function, aiming to intensify the reaction processes with a higher yield of product in a short time, and can be exploited for diverse applications. However, conventional activation strategies such as genetic engineering and chemical modification are generally irreversible for enzyme activity, and they also have many limitations, including complex processes and unpredictable results. Recently, near-infrared (NIR), alternating magnetic field (AMF), microwave and ultrasound irradiation, as real-time and precise activation strategies for enzyme analysis, can address many limitations due to their deep penetrability, sustainability, low invasiveness, and sustainability and have been applied in many fields, such as biomedical and industrial applications and chemical synthesis. These spatiotemporal and controllable activation strategies can transfer light, electromagnetic, or ultrasound energy to enzymes, leading to favorable conformational changes and improving the thermal stability, stereoselectivity, and kinetics of enzymes. Furthermore, the different mechanisms of activation strategies have determined the type of applicable enzymes and manipulated protocol designs that either immobilize enzymes on nanomaterials responsive to light or magnetic fields or directly influence enzymatic properties. To employ these effects to finely and efficiently activate enzyme activity, the physicochemical features of nanomaterials and parameters, including the frequency and intensity of activation methods, must be optimized. Therefore, this review offers a comprehensive overview related to emerging technologies for achieving real-time enzyme activation and summarizes their characteristics and advanced applications.

Herein, a novel laccase gene, Melac13220, was amplified from Methylobacterium extorquens and successfully expressed in Escherichia coli with a molecular weight of approximately 50 kDa. The purified Melac13220 had no absorption peak at 610 nm and remained silent within electron paramagnetic resonance spectra, suggesting that Melac13220 belongs to the non-blue laccase group. Both inductively coupled plasma spectroscopy/optical emission spectrometry (ICP-OES) and isothermal titration calorimetry (ITC) indicated that one molecule of Melac13220 can interact with two iron ions. Furthermore, the optimal temperature of Melac13220 was 65 °C. It also showed a high thermolability, and its half-life at 65 °C was 80 min. Melac13220 showed a very good acid environment tolerance; its optimal pH was 1.5. Cu2+ and Co2+ can slightly increase enzyme activity, whereas Fe2+ could increase Melac13220′s activity five-fold. Differential scanning calorimetry (DSC) indicated that Fe2+ could also stabilize Melac13220. Unlike most laccases, Melac13220 can efficiently decolorize Congo Red and Indigo Carmine dyes even in the absence of a redox mediator. Thus, the non-blue laccase from Methylobacterium extorquens shows potential application value and may be valuable for environmental protection, especially in the degradation of dyes at low pH.

In the present study, the non-blue laccase Melac13220 from Methylobacterium extorquens was immobilized using three methods to overcome problems related to the stability and reusability of the free enzyme: entrapment of the enzyme with sodium alginate, crosslinking of the enzyme with glutaraldehyde and chitosan-, and site-specific covalent immobilization of the enzyme on Fe3O4 nanoparticles by an aldehyde tag. The site-specific covalent immobilization method showed the highest immobilization efficiency and vitality recovery. The optimum temperature of Melac13220 was increased from 65 °C to 80 °C. Immobilized Melac13220 showed significant tolerance to some organic solvents and maintained approximately 80% activity after 10 cycles of use. Differential scanning calorimetry (DSC) indicated that the melting temperature of the enzyme was increased (from 57 °C to 79 °C). Immobilization of Melac13220 also led to improvement in dye decolorization such that Congo Red was completely decolorized within 10 h. The immobilized enzyme can be easily prepared without purification, demonstrating the advantages of using the aldehyde tag strategy and providing a reference for the practical application of different immobilized laccase methods in the industrial field.