SMALPs Are Not Simply Nanodiscs: The Polymer-to-Lipid Ratios of Fractionated SMALPs Underline Their Heterogeneous Nature.

Amphipathic styrene-maleic acid (SMA) copolymers directly solubilize biomembranes into SMA-lipid particles, or SMALPs, that are often regarded as nanodiscs and hailed as a native membrane platform. The promising outlook of SMALPs inspires the discovery of many SMA-like copolymers that also solubilize biomembranes into putative nanodiscs, but a fundamental question remains on how much the SMALPs or SMALP analogues truly resemble the bilayer structure of nanodiscs. This unfortunate ambiguity undermines the utility of SMA or SMA-like copolymers in membrane biology because the structure and function of many membrane proteins depend critically on their surrounding matrices. Here, we report the structural heterogeneity of SMALPs revealed through fractionating SMALPs comprised of lipids and well-defined SMAs via size-exclusion chromatography followed by quantitative determination of the polymer-to-lipid (P/L) stoichiometric ratios in individual fractions. Through the lens of P/L stoichiometric ratios, different self-assembled polymer-lipid nanostructures are inferred, such as polymer-remodeled liposomes, polymer-encased nanodiscs, polymer-lipid mixed micelles, and lipid-doped polymer micellar aggregates. We attribute the structural heterogeneity of SMALPs to the microstructure variations amongst individual polymer chains that give rise to their polydisperse detergency. As an example, we demonstrate that SMAs with a similar S/MA ratio but different chain sizes participate preferentially in different polymer-lipid nanostructures. We further demonstrate that proteorhodopsin, a light-driven proton pump solubilized within the same SMALPs is distributed amongst different self-assembled nanostructures to display different photocycle kinetics. Our discovery challenges the native nanodisc notion of SMALPs or SMALP analogues and highlights the necessity to separate and identify the structurally dissimilar polymer-lipid particles in membrane biology studies.

Multilayer 3D Chiral Folding Polymers and Their Asymmetric Catalytic Assembly.

A novel class of polymers and oligomers of chiral folding chirality has been designed and synthesized, showing structurally compacted triple-column/multiple-layer frameworks. Both uniformed and differentiated aromatic chromophoric units were successfully constructed between naphthyl piers of this framework. Screening monomers, catalysts, and catalytic systems led to the success of asymmetric catalytic Suzuki-Miyaura polycouplings. Enantio- and diastereochemistry were unambiguously determined by X-ray structural analysis and concurrently by comparison with a similar asymmetric induction by the same catalyst in the asymmetric synthesis of a chiral three-layered product. The resulting chiral polymers exhibit intense fluorescence activity in a solid form and solution under specific wavelength irradiation.

Nanodiscs: a versatile nanocarrier platform for cancer diagnosis and treatment.

Cancer therapy is a significant challenge due to insufficient drug delivery to the cancer cells and non-selective killing of healthy cells by most chemotherapy agents. Nano-formulations have shown great promise for targeted drug delivery with improved efficiency. The shape and size of nanocarriers significantly affect their transport inside the body and internalization into the cancer cells. Non-spherical nanoparticles have shown prolonged blood circulation half-lives and higher cellular internalization frequency than spherical ones. Nanodiscs are desirable nano-formulations that demonstrate enhanced anisotropic character and versatile functionalization potential. Here, we review the recent development of theranostic nanodiscs for cancer mitigation ranging from traditional lipid nanodiscs encased by membrane scaffold proteins to newer nanodiscs where either the membrane scaffold proteins or the lipid bilayers themselves are replaced with their synthetic analogues. We first discuss early cancer detection enabled by nanodiscs. We then explain different strategies that have been explored to carry a wide range of payloads for chemotherapy, cancer gene therapy, and cancer vaccines. Finally, we discuss recent progress on organic-inorganic hybrid nanodiscs and polymer nanodiscs that have the potential to overcome the inherent instability problem of lipid nanodiscs.

Asymmetric Catalytic Assembly of Triple-Columned and Multilayered Chiral Folding Polymers Showing Aggregation-Induced Emission (AIE).

Invited for the cover of this issue are Guigen Li's groups at Texas Tech University and Nanjing University. The cover artwork shows that chirality patterns exist from universal to molecular levels showing light emission properties. Read the full story of multilayer 3D chirality and its asymmetric catalytic synthesis at 10.1002/chem.202104102.

Hydrophilic nanoparticles that kill bacteria while sparing mammalian cells reveal the antibiotic role of nanostructures.

To dissect the antibiotic role of nanostructures from chemical moieties belligerent to both bacterial and mammalian cells, here we show the antimicrobial activity and cytotoxicity of nanoparticle-pinched polymer brushes (NPPBs) consisting of chemically inert silica nanospheres of systematically varied diameters covalently grafted with hydrophilic polymer brushes that are non-toxic and non-bactericidal. Assembly of the hydrophilic polymers into nanostructured NPPBs doesn't alter their amicability with mammalian cells, but it incurs a transformation of their antimicrobial potential against bacteria, including clinical multidrug-resistant strains, that depends critically on the nanoparticle sizes. The acquired antimicrobial potency intensifies with small nanoparticles but subsides quickly with large ones. We identify a threshold size (dsilica ~ 50 nm) only beneath which NPPBs remodel bacteria-mimicking membrane into 2D columnar phase, the epitome of membrane pore formation. This study illuminates nanoengineering as a viable approach to develop nanoantibiotics that kill bacteria upon contact yet remain nontoxic when engulfed by mammalian cells.

Asymmetric Catalytic Assembly of Triple-Columned and Multilayered Chiral Folding Polymers Showing Aggregation-Induced Emission (AIE).

Unprecedented chiral multilayer folding 3D polymers have been assembled and regulated by uniform and differentiated aromatic chromophore units between naphthyl piers. Screening catalysts, catalytic systems and monomers were proven to be crucial for asymmetric catalytic Suzuki-Miyaura polycouplings for this assembly. X-ray crystallography of the corresponding dimers and trimers revealed the absolute configuration and the intermolecular packing pattern. Up to 61 960 Mw /41 900 Mn and m/z 4317 for polymers and oligomers, as confirmed by gel permeation chromatography (GPC) and MALDI-TOF MS, indicated that these frameworks were composed of multiple stacked layers. The resulting multiple π-assemblies exhibited remarkable optical properties in aggregated states (photoluminescence in solids and aggregation-induced emission in solutions), as well as reversible redox properties in electrochemical performance.

Polymer-Encased Nanodiscs and Polymer Nanodiscs: New Platforms for Membrane Protein Research and Applications.

Membrane proteins (MPs) are essential to many organisms' major functions. They are notorious for being difficult to isolate and study, and mimicking native conditions for studies in vitro has proved to be a challenge. Lipid nanodiscs are among the most promising platforms for MP reconstitution, but they contain a relatively labile lipid bilayer and their use requires previous protein solubilization in detergent. These limitations have led to the testing of copolymers in new types of nanodisc platforms. Polymer-encased nanodiscs and polymer nanodiscs support functional MPs and address some of the limitations present in other MP reconstitution platforms. In this review, we provide a summary of recent developments in the use of polymers in nanodiscs.

Extraction and reconstitution of membrane proteins into lipid nanodiscs encased by zwitterionic styrene-maleic amide copolymers.

Membrane proteins can be reconstituted in polymer-encased nanodiscs for studies under near-physiological conditions and in the absence of detergents, but traditional styrene-maleic acid copolymers used for this purpose suffer severely from buffer incompatibilities. We have recently introduced zwitterionic styrene-maleic amide copolymers (zSMAs) to overcome this limitation. Here, we compared the extraction and reconstitution of membrane proteins into lipid nanodiscs by a series of zSMAs with different styrene:maleic amide molar ratios, chain sizes, and molecular weight distributions. These copolymers solubilize, stabilize, and support membrane proteins in nanodiscs with different efficiencies depending on both the structure of the copolymers and the membrane proteins.

Environmentally Benign Nanoantibiotics with a Built-in Deactivation Switch Responsive to Natural Habitats.

The massive use of antibiotics in healthcare and agriculture has led to their artificial accumulation in natural habitats, which risks the structure and function of the microbial communities in ecosystems, threatens food and water security, and accelerates the development of resistome. Ideally, antibiotics should remain fully active in clinical services while becoming deactivated rapidly once released into the environment, but none of the current antibiotics meet this criterion. Here, we show a nanoantibiotic design that epitomizes the concept of carrying a built-in "OFF" switch responsive to natural stimuli. The environmentally benign nanoantibiotics consist of cellulose backbones covalently grafted with hydrophilic polymer brushes that by themselves are antimicrobially inactive. In their nanostructured forms in services, these cellulose-based polymer molecular brushes are potent killers for both Gram-positive and Gram-negative bacteria, including clinical multidrug-resistant strains; after services and being discharged into the environment, they are shredded into antimicrobially inactive pieces by cellulases that do not exist in the human body but are abundant in natural habitats. This study illuminates a new concept of mitigating the environmental footprints of antibiotics with rationally designed nanoantibiotics that can be dismantled and disabled by bioorthogonal chemistry occurring exclusively in natural habitats.

Study on Tensile Properties of CFRP Plates under Elevated Temperature Exposure.

Elevated temperature exposure has a negative effect on the performance of the matrix resin in Carbon Fiber Reinforced Plastics (CFRP) plates, whereas limited quantitative research focuses on the deteriorations. Therefore, 30 CFRP specimens were designed and tested under elevated temperatures (10, 30, 50, 70, and 90 °C) to explore the degradations in tensile properties. The effect of temperature on the failure mode, stress-strain curve, tensile strength, elastic modulus and elongation of CFRP plates were investigated. The results showed that elevated temperature exposure significantly changed the failure characteristics. When the exposed temperature increased from 10 °C to 90 °C, the failure mode changed from the global factures in the whole CFRP plate to the successive fractures in carbon fibers. Moreover, with temperatures increasing, tensile strength and elongation of CFRP plates decreases gradually while the elastic modulus shows negligible change. Finally, the results of One-Way Analysis of Variance (ANOVA) show that the degradation of the tensile strength of CFRP plates was due to the impact of elevated temperature exposure, rather than the test error.

Reliability Analysis of Bond Behaviour of CFRP⁻Concrete Interface under Wet⁻Dry Cycles.

Effective bonding between adherents plays a key role in retrofitting concrete structures in civil engineering using fibre-reinforced polymers (FRPs). To ensure structural safety, it is critical to develop design codes, which account for uncertainties of materials, the environment, and load, to estimate bond behaviour under long-term exposure to harsh environments. Therefore, a reliability analysis was performed to study the bond behaviour of FRP⁻concrete interface under wet⁻dry cycles and sustained loading. Thirty double-lap, shear-bonded carbon FRP (CFRP)⁻concrete composite specimens were tested after wet⁻dry cycles and sustained loading exposure. The fracture energy Gf of the bond behavior between CFRP and concrete was directly obtained from the measured local bond-slip curves. Five widely used test methods were adopted to verify the possible distribution types of Gf. Based on the best fit distribution of Gf, a reliability index β was then calculated for the specimens. The effects of wet⁻dry exposure and sustained loading on β were analysed separately. The effects of the mean and standard deviation of the load on β were compared. It was found that the mean had a greater impact on reliability than the standard deviation, but neither changed the regulation of the exponential reduction of β with increasing wet⁻dry cycle time. Their impact was significant for a small number of wet⁻dry cycles but insignificant for more than 4000 wet⁻dry cycles.

Polymer Nanodiscs: Discoidal Amphiphilic Block Copolymer Membranes as a New Platform for Membrane Proteins.

Lipid nanodiscs are playing increasingly important roles in studies of the structure and function of membrane proteins. Development of lipid nanodiscs as a membrane-protein-supporting platform, or a drug targeting and delivery vehicle in general, is undermined by the fluidic and labile nature of lipid bilayers. Here, we report the discovery of polymer nanodiscs, i.e., discoidal amphiphilic block copolymer membrane patches encased within membrane scaffold proteins, as a novel two-dimensional nanomembrane that maintains the advantages of lipid nanodiscs while addressing their weaknesses. Using MsbA, a bacterial ATP-binding cassette transporter as a membrane protein prototype, we show that the protein can be reconstituted into the polymer nanodiscs in an active state. As with lipid nanodiscs, reconstitution of detergent-solubilized MsbA into the polymer nanodiscs significantly enhances its activity. In contrast to lipid nanodiscs that undergo time- and temperature-dependent structural changes, the polymer nanodiscs experience negligible structural evolution under similar environmental stresses, revealing a critically important property for the development of nanodisc-based characterization methodologies or biotechnologies. We expect that the higher mechanical and chemical stability of block copolymer membranes and their chemical versatility for adaptation will open new opportunities for applications built upon diverse membrane protein functions, or involved with drug targeting and delivery.

Polymer-encased nanodiscs with improved buffer compatibility.

Styrene-maleic acid copolymers allow for solubilization and reconstitution of membrane proteins into nanodiscs. These polymer-encased nanodiscs are promising platforms for studies of membrane proteins in a near-physiologic environment without the use of detergents. However, current styrene-maleic acid copolymers display severe limitations in terms of buffer compatibility and ensued flexibility for various applications. Here, we present a new family of styrene-maleic acid copolymers that do not aggregate at low pH or in the presence of polyvalent cations, and can be used to solubilize membrane proteins and produce nanodiscs of controlled sizes.

Hydrophilic Phage-Mimicking Membrane Active Antimicrobials Reveal Nanostructure-Dependent Activity and Selectivity.

The prevalent wisdom on developing membrane active antimicrobials (MAAs) is to seek a delicate, yet unquantified, cationic-hydrophobic balance. Inspired by phages that use nanostructured protein devices to invade bacteria efficiently and selectively, we study here the antibiotic role of nanostructures by designing spherical and rod-like polymer molecular brushes (PMBs) that mimic the two basic structural motifs of bacteriophages. Three model PMBs with different well-defined geometries consisting of multiple, identical copies of densely packed poly(4-vinyl-N-methylpyridine iodide) branches are synthesized by controlled/"living" polymerization, reminiscent of the viral structural motifs comprised of multiple copies of protein subunits. We show that, while the individual linear-chain polymer branch that makes up the PMBs is hydrophilic and a weak antimicrobial, amphiphilicity is not a required antibiotic trait once nanostructures come into play. The nanostructured PMBs induce an unusual topological transition of bacterial but not mammalian membranes to form pores. The sizes and shapes of the nanostructures further help define the antibiotic activity and selectivity of the PMBs against different families of bacteria. This study highlights the importance of nanostructures in the design of MAAs with high activity, low toxicity, and target specificity.

Structure and Dynamics of Extracellular Loops in Human Aquaporin-1 from Solid-State NMR and Molecular Dynamics.

Multiple moderate-resolution crystal structures of human aquaporin-1 have provided a foundation for understanding the molecular mechanism of selective water translocation in human cells. To gain insight into the interfacial structure and dynamics of human aquaporin-1 in a lipid environment, we performed nuclear magnetic resonance (NMR) spectroscopy and molecular dynamics simulations. Using magic angle spinning solid-state NMR, we report a near complete resonance assignment of the human aquaporin-1. Chemical shift analysis of the secondary structure identified pronounced deviations from crystallographic structures in extracellular loops A and C, including the cis Y37-P38 bond in loop A, as well as ordering and immobilization of loop C. Site-specific H/D exchange measurements identify a number of protected nitrogen-bearing side chains and backbone amide groups, involved in stabilizing the loops. A combination of molecular dynamics simulations with NMR-derived restraints and filtering based on solvent accessibility allowed for the determination of a structural model of extracellular loops largely consistent with NMR results. The simulations reveal loop stabilizing interactions that alter the extracellular surface of human AQP1, with possible implications for water transport regulation through the channel. Modulation of water permeation may occur as a result of rearrangement of side chains from loop C in the extracellular vestibule of hAQP1, affecting the aromatic arginine selectivity filter.

A Usual G-Protein-Coupled Receptor in Unusual Membranes.

G-protein-coupled receptors (GPCRs) are the largest family of membrane-bound receptors and constitute about 50% of all known drug targets. They offer great potential for membrane protein nanotechnologies. We report here a charge-interaction-directed reconstitution mechanism that induces spontaneous insertion of bovine rhodopsin, the eukaryotic GPCR, into both lipid- and polymer-based artificial membranes. We reveal a new allosteric mode of rhodopsin activation incurred by the non-biological membranes: the cationic membrane drives a transition from the inactive MI to the activated MII state in the absence of high [H(+)] or negative spontaneous curvature. We attribute this activation to the attractive charge interaction between the membrane surface and the deprotonated Glu134 residue of the rhodopsin-conserved ERY sequence motif that helps break the cytoplasmic "ionic lock". This study unveils a novel design concept of non-biological membranes to reconstitute and harness GPCR functions in synthetic systems.

[Sports injuries and illnesses during the 2008 Olympic Games and 2013 China National Games in Shenyang Division].

OBJECTIVE: To retrospectively analyse the medical imaging examination results of the injuries and illnesses during the 2008 Olympic Games and 2013 China National Games in Shenyang Divison. METHODS: Collected and analyzed the health information and medical imaging examination results from Shengjing Hospital of China Medical University during the two games. RESULTS: There was 9 cases of sports injuries in the 2008 Olympic Games, mainly for knee, ankle ligament injury and muscle sprain, 36 cases of sports injuries in the 2013 China National Games, mainly for head traumas (9 cases), knee injuries (7 cases), ankle injuries (7 cases), shoulder injures (4 cases). Competitions of high risk of being injured were wrestling (10 cases), track and field (8 cases), American football (6 cases). The most common cause of illness were respiratory system (60 cases) and the digestive system (27 cases) in the total 233 cases illnesses in the China National Games. CONCLUSION: Different sports have different characteristic, regularity and mechanism of injury. Medical imaging examination has important value in the diagnosis of injury during large games. The respiratory system and digestive system are the most common illnesses and affect the athletes training and competition as important as injuries. So illness is the focus on the medical care assurance of the large games.

In situ structural studies of Anabaena sensory rhodopsin in the E. coli membrane.

Magic-angle spinning nuclear magnetic resonance is well suited for the study of membrane proteins in the nativelike lipid environment. However, the natural cellular membrane is invariably more complex than the proteoliposomes most often used for solid-state NMR (SSNMR) studies, and differences may affect the structure and dynamics of the proteins under examination. In this work we use SSNMR and other biochemical and biophysical methods to probe the structure of a seven-transmembrane helical photoreceptor, Anabaena sensory rhodopsin (ASR), prepared in the Escherichia coli inner membrane, and compare it to that in a bilayer formed by DMPC/DMPA lipids. We find that ASR is organized into trimers in both environments but forms two-dimensional crystal lattices of different symmetries. It favors hexagonal packing in liposomes, but may form a square lattice in the E. coli membrane. To examine possible changes in structure site-specifically, we perform two- and three-dimensional SSNMR experiments and analyze the differences in chemical shifts and peak intensities. Overall, this analysis reveals that the structure of ASR is largely conserved in the inner membrane of E. coli, with many of the important structural features of rhodopsins previously observed in ASR in proteoliposomes being preserved. Small, site-specific perturbations in protein structure that occur as a result of the membrane changes indicate that the protein can subtly adapt to its environment without large structural rearrangement.

"Frozen" block copolymer nanomembranes with light-driven proton pumping performance.

Cellular membranes are natural nanoengineering devices, where matter transport, information processing, and energy conversion across the nanoscale boundaries are mediated by membrane proteins (MPs). Despite the great potential of MPs for nanotechnologies, their broad utility in engineered systems is limited by the fluidic and often labile nature of MP-supporting membranes. Little is known on how to direct spontaneous reconstitution of MPs into robust synthetic nanomembranes or how to tune MP functions through rational design of these membranes. Here we report that proteorhodopsin (PR), a light-driven proton pump, can be spontaneously reconstituted into "frozen" (i.e., glassy state) amphiphilic block copolymer membranes via a charge-interaction-directed reconstitution mechanism. We show that PR is not enslaved by a fluidic or lipid-based membrane environment. Rather, well-defined block copolymer nanomembranes, with their tunable membrane moduli, act as allosteric regulators to support the structural integrity and function of PR. Versatile membrane designs exist to modulate the conformational energetics of reconstituted MPs, therefore optimizing proteomembrane stability and performance in synthetic systems.