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The use of biological molecules, assemblies and systems in the development of inorganic materials synthesis continues to offer new and exciting alternatives to conventional synthetic strategies. [1] Biological templates, such as protein cages, [2,3] viroid capsules, [4] bacterial rhapidosomes, [5] S-layers, [6] multicellular superstructures, [7] biolipid cylinders, [8,9] and DNA, [10±13] have been utilized to direct the deposition, assembly, and patterning of inorganic nanoparticles and microstructures. In this paper, we report a new approach to the template-directed synthesis of inor-ganic±organic nanotubes using tobacco mosaic virus (TMV).TMV is a remarkably stable virion, remaining intact at temperatures up to 60 C and at pH values between 2 and 10. Each viral particle consists of 2130 identical protein subunits arranged in a helical motif around a single strand of RNA to produce a hollow protein tube, 300´18 nm in size, with a 4 nm-wide central channel. [14,15] The internal and external surfaces of the protein consist of repeated patterns of charged amino acid residues, such as glutamate, aspartate, arginine, and lysine. [16] In principle, these functionalities should offer a wide variety of nucleation sites for surface-controlled inorganic deposition, which, in association with the high thermal and pH stability, could be exploited in the synthesis of unusual materials such as high-aspect-ratio composites and protein-confined inorgan-ic nanowires. Here we show that TMV is a suitable template for reactions such as co-crystallization (CdS and PbS), oxidative hydrolysis (iron oxides), and sol-gel condensation (SiO 2 ) (Fig. 1). Fig. 1. Scheme showing routes for the synthesis of nanotube composites using TMV templates. Clockwise from top right: sol±gel condensation (silica); coprecipitation (PbS and CdS nanocrystals); oxidative hydrolysis (iron oxide).Specific nucleation of CdS on the surface of dispersed particles of TMV was achieved by exposing a buffered suspension of the virions in 10 mM CdCl 2 to H 2 S gas for up to 6 h. Transmission electron microscopy (TEM) revealed the presence of mineralized tubular structures, approximately 50 nm in width, which consisted of a 16 nm thick electrondense outer crust and a 18 nm diameter internal core (Fig. 2a). Energy dispersive X-ray (EDX) analysis confirmed the presence of cadmium and sulfur in individual filaments, and high resolution lattice images indicated that the inorganic coating consisted of disordered aggregates of crystalline CdS particles 5 nm in size (Fig. 2b). The lattice spacings were in agreement with those obtained by selected area electron diffraction (SAED) of individual mineralized tubules, which showed powder patterns (d-spacings 0.336 nm (111), 0.206 nm (220), 0.176 nm (311), 0.133 nm (331), 0.118 nm (422)), corresponding to CdS nanoparticles with the zinc-blende crystal structure. The results were therefore consistent with a relatively uniform coating of CdS nanocrystals on the external surface of the TMV template. Although nucleation of CdS withi...

Methods for organizing materials at the nanometre scale have advanced tremendously in recent years 1,2 . One important objective is the synthesis of patterned arrays of inorganic nanocrystals 3-6 , whose optical, electronic and magnetic properties might find technological uses, for example as memory elements. Techniques such as colloidal crystallization 7-9 , monolayer deposition 10-12 , multilayer casting 13 , molecular crosslinking 14,15 , the use of complementary interactions 16,17 and the synthesis of nanoparticles in patterned etch pits 18 have been used to organize nanocrystals into superlattices. Here we describe the use of bacterial Slayers-self-assembled, two-dimensionally ordered films of proteins that feature in many bacterial cell walls-as templates for the in situ nucleation of ordered two-dimensional arrays of cadmium sulphide nanocrystals about 5 nm in size. Nucleation of the inorganic phase is confined to the pores between subunits in the S-layers. Two-tier stacks of nanoparticles can be formed in the presence of double-layered protein crystals. The structural diversity of S-layers 19,20 , their ease of self-assembly on a wide range of substrates and the potential for surface chemical modification suggest that this approach could be exploited to offer a wide range of ordered nanoparticle arrays.With a few exceptions, S-layers represent an almost universal feature of archaebacterial cell envelopes, and have been identified in many different species of nearly every taxonomic group of walled eubacteria 19,20 . S-layers are two-dimensional crystalline structure of single protein or glycoprotein monomers (relative molecular mass, M r , 40,000 to 200,000), and exhibit either oblique (p1, p2), square (p4) or hexagonal (p3, p6) lattice symmetry with spacings between the morphological units in the range 3-30 nm (ref. 20). Most Slayers are 5-15 nm in thickness, possess pores of identical size and morphology in the 2-6 nm range, and have inner and outer surfaces that are markedly different in topography and physicochemical properties 21 . Although considered primarily as a filtration and structural matrix, S-layers have been implicated in cell wall biomineralization 22,23 , suggesting a possible biomimetic role for these proteins as templates in materials chemistry.Our approach to using S-layer templates for inorganic superlattice construction is illustrated in Fig. 1 (see Methods for details). Figure 2a shows a typical monolayer of the self-assembled S-layer of Bacillus stearothermophilus NRS2004/3a variant 1. The sample was negatively stained with uranyl acetate, which penetrates the nanoporous protein structure to reveal an oblique lattice (space group, p1; a ¼ 9:8 nm, b ¼ 7:5 nm, v ¼ 80Њ). The protein fine structure, which appears white in the stain exclusion pattern, is imaged to a resolution of ϳ2 nm. Similar studies with recrystallized S-layers of B. sphaericus CCM 2177 revealed a square lattice with a unit cell length of 13 nm (data not shown). Mineralization of the exposed inner face of the self-ass...

Double-hydrophilic block copolymers consisting of a long poly(ethylene oxide) block and a short poly(methacrylic acid) block, modified by partial alkylation with dodecylamine (PEO-b-PMAA-C 12 ) were employed as dispersed templates for the controlled precipitation of calcium phosphate from aqueous solution at different pH values. Two new and nontrivial superstructures of an organized inorganic/organic hybrid material were characterized by ultracentrifugation, small-and wide-angle X-ray analysis, and electron microscopy. At pH 3.5 and 4.0, and pH 4.5, 5.0, and 6.3, two different types of discrete nested structures are obtained which consist of hybrid nanofilaments arranged to give an unusual neuronlike morphology. The fibers originate from a core of similar size to the primary polymer aggregates, suggesting that cooperative interactions at a local level between dissolving calcium phosphate clusters and disassembling polymer units could be responsible for the highly anisotropic nature of the secondary growth process.Aging of the nanofilaments grown in acidic solution results in a second hybrid morphology, consisting of compact aggregates with a diameter of about 130 nm, which show the interlocked layer structure of an ordered inorganic/ organic mesophase with a repeat period of about 3 nm. Such calcium phosphate/ polymer nanohybrids with complex morphologies are interesting from the viewpoint of prebiotic structure formation, and might also be useful as novel ceramics precursors, reinforcing fillers, or biomedical implants.

BackgroundUnderstanding the pathogenic role of extracellular vesicles (EVs) in disease and their potential diagnostic and therapeutic utility is extremely reliant on in-depth quantification, measurement and identification of EV sub-populations. Quantification of EVs has presented several challenges, predominantly due to the small size of vesicles such as exosomes and the availability of various technologies to measure nanosized particles, each technology having its own limitations.Materials and MethodsA standardized methodology to measure the concentration of extracellular vesicles (EVs) has been developed and tested. The method is based on measuring the EV concentration as a function of a defined size range. Blood plasma EVs are isolated and purified using size exclusion columns (qEV) and consecutively measured with tunable resistive pulse sensing (TRPS). Six independent research groups measured liposome and EV samples with the aim to evaluate the developed methodology. Each group measured identical samples using up to 5 nanopores with 3 repeat measurements per pore. Descriptive statistics and unsupervised multivariate data analysis with principal component analysis (PCA) were used to evaluate reproducibility across the groups and to explore and visualise possible patterns and outliers in EV and liposome data sets.ResultsPCA revealed good reproducibility within and between laboratories, with few minor outlying samples. Measured mean liposome (not filtered with qEV) and EV (filtered with qEV) concentrations had coefficients of variance of 23.9% and 52.5%, respectively. The increased variance of the EV concentration measurements could be attributed to the use of qEVs and the polydisperse nature of EVs.ConclusionThe results of this study demonstrate the feasibility of this standardized methodology to facilitate comparable and reproducible EV concentration measurements.

The organization of nanostructures across extended length scales is a key challenge in the design of integrated materials with advanced functions. Current approaches tend to be based on physical methods, such as patterning, rather than the spontaneous chemical assembly and transformation of building blocks across multiple length scales. It should be possible to develop a chemistry of organized matter based on emergent processes in which time-and scale-dependent coupling of interactive components generate higher-order architectures with embedded structure. Herein we highlight how the interplay between aggregation and crystallization can give rise to mesoscale selfassembly and cooperative transformation and reorganization of hybrid inorganic-organic building blocks to produce single-crystal mosaics, nanoparticle arrays, and emergent nanostructures with complex form and hierarchy. We propose that similar mesoscale processes are also relevant to models of matrix-mediated nucleation in biomineralization. From the Contents 1. Introduction 2351 2. Kinetic Control of Nucleation and Growth 2352 3. Aggregation-Mediated Pathways of Crystal Growth 2353 4. Mesoscale Self-Assembly of Nanoparticle Arrays 2356 5. Mesoscale Transformations and Emergent Nanostructures 2357 6. Mesoscale Transformations and Matrix-Mediated Nucleation in Biomineralization 2361

Although catalytic mechanisms in natural enzymes are well understood, achieving the diverse palette of reaction chemistries in re-engineered native proteins has proved challenging. Wholesale modification of natural enzymes is potentially compromised by their intrinsic complexity, which often obscures the underlying principles governing biocatalytic efficiency. The maquette approach can circumvent this complexity by combining a robust de novo designed chassis with a design process that avoids atomistic mimicry of natural proteins. Here, we apply this method to the construction of a highly efficient, promiscuous, and thermostable artificial enzyme that catalyzes a diverse array of substrate oxidations coupled to the reduction of H2O2. The maquette exhibits kinetics that match and even surpass those of certain natural peroxidases, retains its activity at elevated temperature and in the presence of organic solvents, and provides a simple platform for interrogating catalytic intermediates common to natural heme-containing enzymes.