Rapid Degradation of SARS-CoV-2 Spike S Protein by A Specific Serine Protease

The S protein of SARS-CoV-2 is a crucial structural and functional component for virus entry. Due to the constant mutation of the virus, there are very limited ways to prevent and control COVID-19. This experiment used a macroscopic SDS-PAGE method and proved that the S protein of wild-type SARS-CoV-2 virus, especially the S1 subunit, is very sensitive to alkaline serine protease with acidic pI (ASPNJ), NJ represents Neanthes japonica (Izuka) from which ASP is purified). ASPNJ cleaves proteins when the carbonyl group of the peptide bond is contributed by arginine or lysine. ASPNJ can degrade the S protein very quickly and effectively in vitro with relative selectivity. It can be inferred that the S, S1 and RBD of SARS-CoV-2 variants can also be easily degraded by ASPNJ. This rapid and strong degradation of the S protein by ASPNJ may become a potential new treatment strategy.     

Light-Induced Dimerization Approaches to Control Cellular Processes

Light-inducible approaches provide a means to control biological systems with spatial and temporal resolution that is unmatched by traditional genetic perturbations. Recent developments of optogenetic and chemo-optogenetic systems for induced proximity in cells facilitate rapid and reversible manipulation of highly dynamic cellular processes and have become valuable tools in diverse biological applications. New expansions of the toolbox facilitate control of signal transduction, genome editing, “painting” patterns of active molecules onto cellular membranes, and light-induced cell cycle control. A combination of light- and chemically induced dimerization approaches have also seen interesting progress. Herein, an overview of optogenetic systems and emerging chemo-optogenetic systems is provided, and recent applications in tackling complex biological problems are discussed.     

Photoreactive DNA as a Tool to Study Replication Protein A Functioning in DNA Replication and Repair

Replication protein A (RPA), eukaryotic single-stranded DNA-binding protein, is a key player in multiple processes of DNA metabolism including DNA replication, recombination and DNA repair. Human RPA composed of subunits of 70-, 32- and 14-kDa binds ssDNA with high affinity and interacts specifically with multiple proteins. The RPA heterotrimer binds ssDNA in several modes, with occlusion lengths of 8–10, 13–22 and 30 nucleotides corresponding to global, transitional and elongated conformations of protein. Varying the structure of photoreactive DNA, the intermediates of different stages of DNA replication or DNA repair were designed and applied to identify positioning of the RPA subunits on the specific DNA structures. Using this approach, RPA interactions with various types of DNA structures attributed to replication and DNA repair intermediates were examined. This review is dedicated to blessed memory of Prof. Alain Favre who contributed to the development of photoreactive nucleotide derivatives and their application for the study of protein–nucleic acids interactions.     

Tyrosyl-tRNA Synthetase: A Housekeeping Protein and an Attractive Harbinger of Cellular Death

Cytokine-type activities are observed for the human tyrosyl-tRNA synthetase, largely considered as an essential enzyme for protein synthesis, only after cleavage into two fragments. These peptide fragments are novel elements in the orchestration of the tissue response to a cellular suicide program and should be viewed as highly differentiated adaptions of peptide modules with biological activity in more than one kind of environment.     

Small‐Molecule PROTACS: New Approaches to Protein Degradation

The current inhibitor‐based approach to therapeutics has inherent limitations owing to its occupancy‐based model: 1) there is a need to maintain high systemic exposure to ensure sufficient in vivo inhibition, 2) high in vivo concentrations bring potential for off‐target side effects, and 3) there is a need to bind to an active site, thus limiting the drug target space. As an alternative, induced protein degradation lacks these limitations. Based on an event‐driven model, this approach offers a novel catalytic mechanism to irreversibly inhibit protein function by targeting protein destruction through recruitment to the cellular quality control machinery. Prior protein degrading strategies have lacked therapeutic potential. However, recent reports of small‐molecule‐based proteolysis‐targeting chimeras (PROTACs) have demonstrated that this technology can effectively decrease the cellular levels of several protein classes.     

Accelerated Degradation of the D2 Protein of Photosystem II Under Ultraviolet Radiation

The D2 protein of photosystem II is relatively stable in vivo under photosynthetic active radiation, but its degradation accelerates under UVB radiation. Little is known about accelerated D2 protein degradation. We characterized wavelength dependence and sensitivity toward photosystem II inhibitors. The in vivo D2 degradation spectrum resembles the pattern for the rapidly turning over D1 protein of photosystem II, with rates being maximal in the UVB region. We propose that D2 degradation, like D1 degradation, is activated by distinct photosensitizers in the UVB and visible regions of the spectrum. In both wavelength regions, photosystem II inhibitors that are known to be targeted to the D1 protein affect D2 degradation. This suggests that degradation of the two proteins is coupled, D2 degradation being influenced by events occurring at the Q_B niche on the D1 protein.     

Pressure—A Gateway to Fundamental Insights into Protein Solvation,Dynamics, and Function

Now that the centennial anniversary of the first report on pressure denaturation of proteins by Nobel Laureate P. W. Bridgman can be celebrated, this Review on the application of high pressure as a key variable for studying the energetics and interactions of proteins appears. We demonstrate that combined temperature–pressure‐dependent studies help delineate the free‐energy landscape of proteins and elucidate which features are essential in determining their stability. Pressure perturbation also serves as an important tool to explore fluctuations in proteins and reveal their conformational substates. From shaping the free‐energy landscape of proteins themselves to that of their interactions, conformational fluctuations not only dictate a plethora of biological processes, but are also implicated in a number of debilitating diseases. Finally, the advantages of using pressure to explore biomolecular assemblies and modulate enzymatic reactions are discussed.     

Protein Oligomer Engineering: A New Frontier for Studying Protein Structure,Function, and Toxicity

Prevalent in nature, protein oligomers play critical roles both physiologically and pathologically. The multimeric nature and conformational transiency of protein oligomers greatly complicate a more detailed glimpse into the molecular structure as well as function. In this minireview, the oligomers are classified and described on the basis of biological function, toxicity, and application. We also define the bottlenecks in recent oligomer studies and further review numerous frontier methods for engineering protein oligomers. Progress is being made on many fronts for a wide variety of applications, and protein grafting is highlighted as a promising and robust method for oligomer engineering. These advances collectively allow the engineering and design of stabilized oligomers that bring us one step closer to understanding their biological functions, toxicity, and a wide range of applications.     

Aptamer Inhibits Tumor Growth by Leveraging Cellular Proteasomal Degradation System to Degrade c-Met in Mice

Current action mechanisms for aptamer-based therapeutics depend on occupancy-driven pharmacology to mediate protein functions. We report a new mechanism where aptamers leverage cellular proteasomal degradation system to degrade proteins for cancer treatment. A DNA aptamer (hereinafter referred to as c-Met-Ap) binds to the extracellular domain of mesenchymal-epithelial transition factor (c-Met) and selectively induces c-Met phosphorylation at Y1003 and Y1349. The phosphorylation of Y1003 recruits E3 ubiquitin ligase casitas B-lineage lymphoma, causing c-Met ubiquitination and degradation in the proteasome. Furthermore, c-Met-Ap can induce a decrease in the heterodimeric partner proteins of c-Met and the downstream effector proteins in the c-Met signal axis, effectively inhibiting tumor growth in A549 tumor-bearing BALB/c mice. Our study uncovers a novel, actionable mechanism for aptamer therapeutics and opens a new avenue for developing highly efficient anticancer drugs.     

Protein Functionalization Revised: N‐tert‐butoxycarbonylation as an Elegant Tool to Circumvent Protein Crosslinking

The protection of primary amines available in proteins holds great potential to introduce a plethora of diverse functionalities along the protein backbone (e.g., via its carboxylic acid or alcohol moieties) while circumventing the crosslinking issue using conventional approaches. This paper reports on a straightforward and efficient proof‐of‐concept including the chemoselective N‐tert‐butyloxycarbonylation of the primary amines in the protein gelatin (gel‐NH‐BOC), followed by introducing crosslinkable methacrylamide moieties. The reaction is performed successfully under relatively mild conditions (50 °C). Following selective protein functionalization, the deprotection is realized by adding a catalytic amount of an aqueous hydrogen chloride solution. The present communication illustrates the occurrence of a straightforward and selective deprotection procedure, which is typically required to circumvent the occurrence of acidic hydrolysis of the protein backbone. The results hold promise for a large range of biomedical applications in which the presence of primary amines is essential for preserving the biological activity.

     

pH-Responsive Side Chains as a Tool to Control Aqueous Self-Assembly Mechanisms

pH-Tunable nanoscale morphology and self-assembly mechanism of a series of oligo(p-phenyleneethynylene) (OPE)-based bolaamphiphiles featuring poly(ethylene imine) (PEI) side chains of different length and degree of hydrolysis are described. Protonation and deprotonation of the PEI chains by changing the pH alters the hydrophilic/hydrophobic balance of the systems and, in turn, the strength of intermolecular interactions between the hydrophobic OPE moieties. Low pH values (3) lead to weak interaction between the OPEs and result in spherical nanoparticles, in which aggregation follows an isodesmic mechanism. In contrast, higher pH values (11) induce deprotonation of the polymer chains and lead to a stronger, cooperative aggregation into anisotropic nanostructures. Our results demonstrate that pH-responsive chains can be exploited as a tool to tune self-assembly mechanisms, which opens exciting possibilities to develop new stimuli-responsive materials.     

Sonochemical Degradation of Gelatin Methacryloyl to Control Viscoelasticity for Inkjet Bioprinting

Inkjet printing enables the mimicry of the microenvironment of natural complex tissues by patterning cells and hydrogels at a high resolution. However, the polymer content of an inkjet-printable bioink is limited as it leads to strong viscoelasticity in the inkjet nozzle. Here it is demonstrated that sonochemical treatment controls the viscoelasticity of a gelatin methacryloyl (GelMA) based bioink by shortening the length of polymer chains without causing chemical destruction of the methacryloyl groups. The rheological properties of treated GelMA inks are evaluated by a piezo-axial vibrator over a wide range of frequencies between 10 and 10 000 Hz. This approach enables to effectively increase the maximum printable polymer concentration from 3% to 10%. Then it is studied how the sonochemical treatment effectively controls the microstructure and mechanical properties of GelMA hydrogel constructs after crosslinking while maintaining its fluid properties within the printable range. The control of mechanical properties of GelMA hydrogels can lead fibroblasts more spreading on the hydrogels. A 3D cell-laden multilayered hydrogel constructs containing layers with different physical properties is fabrictated by using high-resolution inkjet printing. The sonochemical treatment delivers a new path to inkjet bioprinting to build microarchitectures with various physical properties by expanding the range of applicable bioinks.     

Absorption and emission spectroscopic characterisation of the LOV2-His domain of phot from Chlamydomonas reinhardtii

The absorption and emission behaviour of flavin mononucleotide (FMN) in the wild-type light, oxygen and voltage sensitive domain LOV2 of the photoreceptor phot from the green alga Chlamydomonas reinhardtii is studied. Actually a LOV2-His protein (LOV2 domain bound at N-terminal to 15 His aminoacids via a Gly aminoacid) expressed in an Escherichia coli strain is investigated. For fresh samples stored in the dark an initial fluorescence quantum yield of ϕ_F = 0.12 ± 0.01 and an effective fluorescence lifetime of τ_F = 2.4 ± 0.1 ns are determined. Blue-light photo-excitation generates an intermediate photoproduct (flavin-C(4a)-cysteinyl adduct with absorption peak at 390 nm) resulting in an intensity-dependent fluorescence quenching. In the aqueous solutions at pH 8 approximately 3.8% of the FMN molecules are not bound to the protein binding pocket, whereas 96.2% are non-covalently bound. Even at high-intensity light excitation at 428 nm a fraction of about 7% of the non-covalently bound FMN remains non-converted to an FMN-Cys adduct because of photo-induced back-relaxation of the adduct to non-covalently bound FMN. Two holo-LOV2-His conformations with different adduct recovery time constants are revealed by spectrally and temporally resolved fluorescence and absorption measurements: A fraction of about 48% forms FMN-Cys adducts with a fast recovery time constant of τ_Ad,f = 19 ± 2 s in the dark, and the rest forms adducts with a slow recovery time constant of τ_F,s = 5.5 ± 1 min. Prolonged blue light irradiation of the flavin-C(4a)-cysteinyl adducts reduces their ability to recover back in the dark to non-covalently bound FMN (photo-induced permanent adduct formation). Numerical simulations of the intensity-dependent absorption depletion reveals a quantum yield of intermediate photo-adduct formation of ϕ_Ad = 0.9 ± 0.1. Simulation of the adduct absorption dynamics gives a quantum yield of photo-induced adduct back-relaxation of ϕ_Ad,b = 0.15 ± 0.01 and a quantum yield of photo-induced permanent adduct formation of ϕ_Ad,p = (2.6 ± 0.5) × 10⁻⁴.     

TiO2/BixTiyOz气相光催化降解苯的动力学模型及反应机理

在密闭不锈钢反应器内考察了TiO2/BixTjyOz催化剂气相光催化降解苯的性能.结果表明,TiO2负载于Bi12TiO20,Bi2Ti2O7和Bi4Ti3O12上制成的催化剂,光催化活性得到很人的提高,TiO2最佳负载量为2.0%;其中,TiO2/Bi12TiO20的光催化活性最高,苯最高转化率是纯TiO2的2倍,催化剂使用寿命也延长了1倍.在本文实验条件下,TiO2/Bi12TiO20上苯气相光催化降解符合Lang-muir-Hinshelwood动力学模型,光催化反应速率常数k和Langmuir吸附常数K分别为0.006 4mg/(L·min)和9.670 2L/mg.采用红外光谱对失活的催化剂进行表征,结果表明催化剂表面出现了羰基与羟基等的振动峰,同时检测到主要的中间产物是2,6-二叔丁基-4-甲基苯酚,它吸附在催化剂表面活性化上而导致催化剂失活.最后推测了苯在催化剂表面气相光催化降解的反应机理.