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In addition to nucleic acids, a variety of other biomolecules have also been found on the plasma membrane. Although researchers have realized that RNA has the ability to bind to membrane vesicles in vitro, little is known about whether and how RNA connects to the plasma membrane of the cell. The combination of high-throughput sequencing and in situ labeling methods provides an innovative approach for large-scale identification of subcellular RNAs. Here, we applied the recently published method APEX-seq and identified 75 RNAs related to the plasma membrane, in which lncRNA PMAR72 (plasma membrane-associated RNA AL121772.1) has a considerable affinity with sphingomyelin (SM) and localizes within distinct membrane foci. Our findings will provide some new evidence to elaborate the relationship between RNA and the plasma membrane of mammalian cells. [ABSTRACT FROM AUTHOR]

Arrestins recognize different receptor phosphorylation patterns and convert this information to selective arrestin functions to expand the functional diversity of the G protein-coupled receptor (GPCR) superfamilies. However, the principles governing arrestin-phospho-receptor interactions, as well as the contribution of each single phospho-interaction to selective arrestin structural and functional states, are undefined. Here, we determined the crystal structures of arrestin2 in complex with four different phosphopeptides derived from the vasopressin receptor-2 (V2R) C-tail. A comparison of these four crystal structures with previously solved Arrestin2 structures demonstrated that a single phospho-interaction change results in measurable conformational changes at remote sites in the complex. This conformational bias introduced by specific phosphorylation patterns was further inspected by FRET and 1H NMR spectrum analysis facilitated via genetic code expansion. Moreover, an interdependent phospho-binding mechanism of phospho-receptor-arrestin interactions between different phospho-interaction sites was unexpectedly revealed. Taken together, our results provide evidence showing that phospho-interaction changes at different arrestin sites can elicit changes in affinity and structural states at remote sites, which correlate with selective arrestin functions. The interaction between a GPCR, such as the vasopressin receptor-2 (V2R), and arrestin depends on the receptors' phosphorylation pattern. Here authors use FRET and NMR to analyze the phosphorylation patterns of the V2R-arrestin complex and show that phospho-interactions are the key determinants of selective arrestin conformational states and correlated functions. [ABSTRACT FROM AUTHOR]

Visible light-driven redox reactions have been widely adopted for the production of chemicals to combat energy shortage and global warming. Key elements of such a reaction system include a photosensitizer, a catalyst, and an electron source. In this review, we introduce the small molecules and nanoparticles that are widely used as photosensitizers, as well as the development of a photosensitizer protein that is based on the expansion of genetic code, with a fluorescent protein that is used as a scaffold. Visible light-driven enzymes using proteins as photosensitizers or as catalysts such as carbon monoxide dehydrogenase (CODH), formic acid dehydrogenase (FDH), hydrogenase, nitrogenase, cytochrome P450 BM3, and alkane synthase are then described. CODH can be coupled with photosensitizing nanoparticles to reduce CO2 to CO, and hydrogenase can produce H2 using high-energy electrons produced from dye-sensitized nanoparticles. When water-soluble zinc porphyrin is coupled with FDH, visible light drives CO2 to produce formic acid. Nitrogenase can reduce N2 to NH3 using CdS nanoparticle as photosensitizer. Cytochrome P450 BM3 can be enhanced by a visible light-driven redox system and thus by hydroxylate lauric acid or fatty acids. CvFAP, an alkane synthase, can decarboxylate palmitic acid to pentadecane under blue light excitation. Moreover, we describe a genetically encoded photosensitive protein, which mimics the function of natural photosynthesis and catalyzes the conversion of CO2 to CO when covalently attached with a Ni–terpyridine complex. [ABSTRACT FROM AUTHOR]

Abstract: Fluorescent proteins (FPs) with emission wavelengths in the far-red and infrared regions of the spectrum provide powerful tools for deep-tissue and super-resolution imaging. The development of red-shifted FPs has evoked widespread interest and continuous engineering efforts. In this article, based on a computational design and genetic code expansion, we report a rational approach to significantly expand and red-shift the chromophore of green fluorescent protein (GFP). We applied computational calculations to predict the excitation and emission wavelengths of a FP chromophore harboring unnatural amino acids (UAA) and identify in silico an appropriate UAA, 2-amino-3-(6-hydroxynaphthalen-2-yl)propanoic acid (naphthol-Ala). Our methodology allowed us to formulate a GFP variant (cpsfGFP-66-Naphthol-Ala) with red-shifted absorbance and emission spectral maxima exceeding 60 and 130 nm, respectively, compared to those of GFP. The GFP chromophore is formed through autocatalytic post-translational modification to generate a planar 4-(p-hydroxybenzylidene)-5-imidazolinone chromophore. We solved the crystal structure of cpsfGFP-66-naphthol-Ala at 1.3 Å resolution and demonstrated the formation of a much larger conjugated π-system when the phenol group is replaced by naphthol. These results explain the significant red-shifting of the excitation and emission spectra of cpsfGFP-66-naphthol-Ala.Graphical abstract: [ABSTRACT FROM AUTHOR]

Metalloproteins have inspired chemists for many years to synthesize artificial catalysts that mimic native enzymes. As a complementary approach to studying native enzymes or making synthetic models, biosynthetic approach using small and stable proteins to model native enzymes has offered advantages of incorporating non-covalent secondary sphere interactions under physiological conditions. However, most biosynthetic models are restricted to natural amino acids. To overcome this limitation, incorporating unnatural amino acids into the biosynthetic models has shown promises. In this review, we summarize first synthetic, semisynthetic and biological methods of incorporates unnatural amino acids (UAAs) into proteins, followed by progress made in incorporating UAAs into both native metalloproteins and their biosynthetic models to fine-tune functional properties beyond native enzymes or their variants containing natural amino acids, such as reduction potentials of azurin, O reduction rates and percentages of product formation of HCO models in Mb, the rate of radical transport in ribonucleotide reductase (RNR) and the proton and electron transfer pathways in photosystem II (PS II). We also discuss how this endeavour has allowed systematic investigations of precise roles of conserved residues in metalloproteins, such as Met121 in azurin, Tyr244 that is cross-linked to one of the three His ligands to CuB in HCO, Tyr122, 356, 730 and 731 in RNR and TyrZ in PS II. These examples have demonstrated that incorporating UAAs has provided a new dimension in our efforts to mimic native enzymes and in providing deeper insights into structural features responsible high enzymatic activity and reaction mechanisms, making it possible to design highly efficient artificial catalysts with similar or even higher activity than native enzymes. [ABSTRACT FROM AUTHOR]

Rational protein design is a powerful strategy, not only for revealing the structure and function relationship of natural metallo-proteins, but also for creating artificial metalloproteins with improved properties and functions. Myoglobin (Mb), a small heme protein created by nature with diverse functions, has been shown to be an ideal scaffold for rational protein design. The progress reviewed herein includes fine-tuning its native functions of O binding and transport, peroxidase activity and nitrite reductase (NIR) activity, and rational expanding its functionalities to peroxygenase, heme-copper oxidase (HCO), nitric oxide reductase (NOR), as well as hydroxylamine reductase. These studies have enhanced our understanding of how metalloproteins work in nature, and provided insights for rational design of functional metalloproteins for practical applications in the future. [ABSTRACT FROM AUTHOR]

We consider a technology for granulation of asphalt from its mixture with supercritical pentane and construct a mathematical model for the process. We compare two nozzle designs intended for spraying the mixture and flash boiling of pentane. We show that a nozzle with a conical section is not susceptible to clogging by the asphalt. The pentane vaporization rate and the fraction of vapor phase increase owing to the presence of the asphalt phase. [ABSTRACT FROM AUTHOR]

The modeling and simulation (M&S) architecture describes and defines the relationship between the different parts of a simulation. The simulation system architecture and simulation support platform architecture are discussed separately. The simulation support platform architecture consists of the management layer, the resource layer, the communication layer, the application layer and the infrastructure layer. The best way is to design and realize the M&S collaborative environment for simulation support platform in the resource-communication-application three-dimension space. [ABSTRACT FROM AUTHOR]