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The role of copper in cancer progression has been established since decades. Additionally, copper is able to stimulate angiogenesis through the control of VEGF expression and activity in endothelial cells. In this paper a tetrapeptide, belonging to the histidine-proline-rich glycoprotein (HPRG) and encompassing four repeats of the sequence GHHPH (named TetraHPRG), was synthesized and its copper(ii) complex species were characterized by means of potentiometry, UV-vis, circular dichroism (CD), electron paramagnetic resonance (EPR) and electron spray ionization mass spectrometry (ESI-MS). Moreover, a peptide covalently bound through an amidic bond to trehalose (TH-TetraHPRG) was designed and synthesized as a prodrug system. The activity of both TetraHPRG and TH-TetraHPRG molecules on copper and VEGF induced angiogenic responses in endothelial cells was assessed. The two peptides show a similar and effective anti-angiogenic activity on both molecular and cellular responses. Since the trehalose derivative has a higher resistance to enzymatic degradation, it can be further exploited as a potential drug delivery system with anti-angiogenic activity.

In this paper, we present a coarse replica exchange molecular dynamics (REMD) approach, based on kinetic Monte Carlo (kMC). The new development significantly can reduce the amount of replicas and the computational cost needed to enhance sampling in protein simulations. We introduce 2 different methods which primarily differ in the exchange scheme between the parallel ensembles. We apply this approach on folding of 2 different β-stranded peptides: the C-terminal β-hairpin fragment of GB1 and TrpZip4. Additionally, we use the new simulation technique to study the folding of TrpCage, a small fast folding α-helical peptide. Subsequently, we apply the new methodology on conformation changes in signaling of the light-oxygen voltage (LOV) sensitive domain from Avena sativa (AsLOV2). Our results agree well with data reported in the literature. In simulations of dialanine, we compare the statistical sampling of the 2 techniques with conventional REMD and analyze their performance. The new techniques can reduce the computational cost of REMD significantly and can be used in enhanced sampling simulations of biomolecules.

High blood pressure is considered as a significant health problem worldwide. In addition to numerous preventive and therapeutic drug treatments, important advances have been achieved in the identification of dietary compounds that may contribute to cardiovascular health. Among these compounds, peptides with antihypertensive properties received special attention in the past 15 years. Although milk proteins are still the main source of antihypertensive peptides, recently a remarkable increase has been noticed in the report of antihypertensive peptides released from other dietary sources. Most of these peptides have demonstrated their properties by in vitro assays. However, the evidence for their beneficial antihypertensive effects has to be based on animal experiments and clinical trials. This paper reviews the current data of the blood pressure-lowering activity of food-derived peptides demonstrated in vivo (animal models and humans). Other aspects, such as the mechanism of action and bioavailability of these peptides which play a key role in their antihypertensive effects are also summarized in this review., (This journal is © The Royal Society of Chemistry 2012)

This paper describes a methodology that correlates experimental chemical shifts (at the alpha proton) of proteins with their geometrical data (both dihedral angles and distances) obtained from 13 representative proteins, which are taken from the Protein Data Bank (PDB) and the BioMagRes Data Bank (BMRB). To this end, the experimentally measured proton chemical shifts of simple amides have been correlated with DFT-based calculated structures, at the B3PW91/6-31G* level. This results in a series of mathematical relationships, which are extrapolated to the above-mentioned proteins giving rise to a modified equation for such skeleta. It is relevant to note that the equation is also supported by a clear comparison with NMR data of a protein beyond the chosen set, such as insulin, even with lower errors. The model also relates the dependence of chemical shifts on hydrophobic and anisotropic effects at the amino acid residues.

Substances that mediate the import of proteins into cells, "carriers", have many potential applications. The most potentially useful carriers do not have to be covalently linked to their protein cargoes. However, a common problem with all carrier molecules is that they tend to deposit the cargo proteins into endosomes; diffuse distribution in the cytosol is the desired outcome. This paper describes the import of four different labeled (Alexa Fluor 488) proteins (avidin, recombinant streptavidin, bovine serum albumin, and beta-galactosidase), with the well-known non-covalent carrier called pep-1 (also known as Chariot), with R(8) (a molecule that is not widely appreciated to import protein cargoes via a non-covalent mode of action), and with a new molecule called azo-R(8). The data collected from fluorescence microscopy and flow cytometry indicate that all three non-covalent carriers can facilitate transport. At 37 degrees C, import into endocytic compartments dominates, but at 4 degrees C weak, diffuse fluorescence is observed in the cytosol, indicative of a favorable mode of action.

Protein S-acylation is an important lipid modification characteristic for heterogeneity in the acyl chain and dynamicity in the acylation/deacylation cycle. Most S-acylproteomic research has been limited by indirect identification of modified proteins/peptides without attached fatty acids, resulting in the failure to precisely characterize S-acylated sites with attached fatty acids. The study of S-acylation turnover is still limited at the protein level. Herein, aiming to site-specifically profile both the heterogeneity and the turnover of S-acylation, we first developed a site-specific strategy for intact S-acylated peptide analysis by introducing an acid cleavable bioorthogonal tag into a metabolic labelling method (ssMLCC). The cleavable bioorthogonal tag allowed for the selective enrichment and efficient MS analysis of intact S-acylated peptides so that S-acylated sites and their attached fatty acids could be directly analysed, enabling the precise mapping of S-acylated sites, as well as circumventing false positives from previous studies. Moreover, 606 S-palmitoylated (C16:0) sites of 441 proteins in HeLa cells were identified. All types of S-acylated peptides were further characterized by an open search, providing site-specific profiling of acyl chain heterogeneity, including S-myristoylation, S-palmitoylation, S-palmitoleylation, and S-oleylation. Furthermore, site-specific monitoring of S-palmitoylation turnover was achieved by coupling with pulse-chase methods, facilitating the detailed observation of the dynamic event at each site in multi-palmitoylated proteins, and 85 rapidly cycling palmitoylated sites in 79 proteins were identified. This study provided a strategy for the precise and comprehensive analysis of protein S-acylation based on intact S-acylated peptide analysis, contributing to the further understanding of its complexity and biological functions. [ABSTRACT FROM AUTHOR]

The evolution of the SARS-CoV2 coronavirus spike S-protein is studied using a mass spectrometry based protein phylogenetic approach. A study of a large dataset comprising sets of peptide masses derived from over 3000 proteins of the SARS-CoV2 virus shows that the approach is capable of resolving and correctly displaying the evolution of the major variants of concern. Using these numerical datasets, through a pairwise comparison of sets of proteolytic peptide masses for each protein, the tree is built without the need for the sequence data itself or any sequence alignment. In the same analysis, single point mutations are calculated from peptide mass differences of different protein sets and these are displayed at the branch nodes on the tree. The tree topology is found to be consistent with that generated using conventional sequence-based phylogenetics by a manual visualisation and using a tree comparison algorithm. The mass tree resolves major variants of the virus and displays non-synonymous mutations, calculated based on the mass data alone, on the tree that enable protein evolution to be charted and tracked along interconnected branches. Tracking the evolution of the SARS-CoV2 coronavirus S-protein is of particular importance given its role in the attachment of the virus to host cells ahead of viral replication. [ABSTRACT FROM AUTHOR]

Biothiols play essential roles in balancing the redox state and modulating cellular functions. Fluorescent probes for monitoring/labelling biothiols often suffer from slow reaction rates, strong background fluorescence and cytotoxic byproduct release. Thus, developing facile and versatile probes to overcome the challenges is still in high demand. Here, we report four coumarin–maleimides as fast responding and fluorogenic probes to detect GSH or label peptides/proteins. The probes quantitatively and selectively react with GSH via Michael addition within 1–2 min, achieving an 11–196-fold increase in fluorescence quantum yield via blockage of the photoinduced electron transfer (PET) process. Optimized probe 4 is applied for the detection of GSH in vitro (A549 cells) and in vivo (zebrafish embryos). Taking advantage of the fast Michael addition between the maleimide moiety and the sulfhydryl group, we expand the application of our method for fluorescent labelling of peptides/proteins and for tracking their cellular uptake process. The labelling strategy works for both Cys-bearing and Cys-free proteins after the introduction of a sulfhydryl group using Traut's reagent. Fluorescence assay reveals that the TAT-peptide can efficiently enter cells, but H3 protein, part of nucleosomes, prefers to bind on the cell membrane by electrostatic interactions, shedding light on the cellular uptake activity of nucleosomes and affording a potential membrane staining strategy. Overall, our study illustrates the broad potential of coumarin–maleimide based dual-functional probes for GSH detection and versatile protein labelling in biochemical research. [ABSTRACT FROM AUTHOR]

Dirhodium complexes of general formula [Rh2(O2CR)4]L2 are a well-known class of bimetallic compounds that are used as efficient catalysts for a variety of reactions and have been shown to be potent antibacterial and anticancer agents. The catalytic and biological properties of these complexes largely depend on the nature of the bridging carboxylate ligands. Trifluoroacetate (tfa)-containing dirhodium compounds have been used to build artificial metalloenzymes upon reaction with peptides and have been shown to be more cytotoxic than dirhodium tetraacetate. However, there is no structural information on the interaction between these compounds and proteins. Here, cis-Rh2(μ-O2CCH3)2(μ-O2CCF3)2 ([cis-Rh2(OAc)2(tfa)2]) has been synthesized and its reaction with bovine pancreatic ribonuclease (RNase A) and hen egg white lysozyme (HEWL) was analyzed using a combination of different techniques, including Fluorine-19 nuclear magnetic resonance spectroscopy and macromolecular X-ray crystallography, with the aim to unveil the differences in the reactivity of tfa-containing dihrodium complexes with proteins when compared to [Rh2(OAc)4]. [cis-Rh2(OAc)2(tfa)2] and [Rh2(OAc)4] bind the N atoms of His side chains of RNase A at the axial position; however the fluorine-containing compound rapidly loses its tfa ligands, while [Rh2(OAc)4] can retain the acetate ligands upon protein binding. The reactivity of [cis-Rh2(OAc)2(tfa)2] with HEWL is slightly distinct when compared to that of [Rh2(OAc)4] under the same experimental conditions; however, both [cis-Rh2(OAc)2(tfa)2] and [Rh2(OAc)4] degrade when soaked within HEWL crystals. These results provide a structural-based guide for the design of new heterogenous chiral dirhodium/peptide and dirhodium/protein adducts with application in the fields of organic synthesis and asymmetric catalysis. [ABSTRACT FROM AUTHOR]

All stakeholders in cultural heritage share an interest in fabrication methods and material technology. Until now methods for analysis of organic materials, particularly proteins, have not been widely available to researchers at cultural institutions. This paper will describe an analytical method for the identification of collagen-based materials from soft tissue sources and show examples of its application to diverse museum objects. The method, peptide mass fingerprinting (PMF), uses enzymatic digestion of extracted proteins to produce a mixture of peptides. The mass spectrum of the mixture contains characteristic marker ions―a peptide mass fingerprint―which are compared to species-specific markers from references as the basis of identification. Preliminary results indicate that analysis of materials from aged samples, several different tissue types, and tanned or untanned materials yields comparable PMF results. Significantly, PMF is simple, rapid, sensitive and specific, has been implemented in a museum laboratory, and is being practiced successfully by non-specialists. [ABSTRACT FROM AUTHOR]

Milk is a natural suspension of essential nutrients and can be exploited not only as a beverage consumed directly or transformed into a range of traditional dairy foods, but also as a source of ingredients for food product innovation. In addition, by-products from the dairy industry, which used to constitute an economical and environmental problem, are still rich in proteins, lactose and minerals. Therefore, from a green point of view, they can be valorized and are being exploited for multiple food and non-food applications. This paper aims to give an overview of the different applications of milk ingredients and by-products from the dairy industry. Additionally, enzymatic modification and production of bioactive peptides and bioemulsifiers broaden their functionality and applicability. Finally, a critical review of the most promising aspects as well as some features that need further development is presented. The unmet needs, the cross fertilization in between protein domains, the carbon footprint requirements, the environmental necessities, the health and wellness new demands, etc. are dominant factors in the search for an innovation approach outlining the potential that these “apparent” constraints oblige science and technology to account for. The adjacent technology analysis opportunity that such a review generates is, in essence, an open innovation step. [ABSTRACT FROM AUTHOR]

The accurate determination of disulfide bonds for protein identification is in high demand. In this study, a simple electrochemical-mass spectrometry (EC-MS) method that possesses advantages of real-time information, simultaneous disulfide bond electroreduction and tagging was developed. In this EC-MS, an ITO glass corner functions as a counter electrode and spray system, and allows the direct sampling of the droplet-scale reacting solution in real-time. The application of this method was successfully demonstrated by electrochemical reduction of oxidized glutathione (GSSG) with one disulfide bond as well as insulin with multiple disulfide bonds. The preferred electroreduction of intermolecular-bonded disulfides for insulin has been observed and the intramolecular bond was not favored. Moreover, simultaneously tagging the formed thiol residues from electroreduction of GSSG using electrogenerated intermediates such as dopamine orthoquinone (DQ) and benzoquinone (Q) was performed. A proof-of-concept was also demonstrated with a large molecule, β-lactoglobulin A. The relationship between signal strength and operating parameters was also studied. This method successfully detected the reduction reaction of the disulfide bond in the polypeptide and protein. The detection limit (S/N ≥ 3) is 0.398 μg mL−1. These results suggest that this EC-MS platform can count cysteine moieties in proteins using a single drop of sample and in real-time and is promising for protein identification experiments. [ABSTRACT FROM AUTHOR]

A split intein-based method has been developed to detect peptide:N-glycanase (PNGase) activity in live cells. PNGase cleaves the linkage between N,N′-diacetylchitobiose and the Asn side-chain of N-intein peptides and the products react rapidly with C-intein by protein trans-splicing to generate an active luciferase. [ABSTRACT FROM AUTHOR]

Ion mobility mass spectrometry experiments enable the characterization of mass, assembly, and shape of biological molecules and assemblies. Here, a new radio-frequency confining drift cell is characterized and used to measure the mobilities of peptide, protein, and protein complex ions. The new drift cell replaced the traveling-wave ion mobility cell in a Waters Synapt G2 HDMS. Methods for operating the drift cell and determining collision cross section values using this experimental set up are presented within the context of the original instrument control software. Collision cross sections for 349 cations and anions are reported, 155 of which are for ions that have not been characterized previously using ion mobility. The values for the remaining ions are similar to those determined using a previous radio-frequency confining drift cell and drift tubes without radial confinement. Using this device under 2 Torr of helium gas and an optimized drift voltage, denatured and native-like ions exhibited average apparent resolving powers of 14.2 and 16.5, respectively. For ions with high mobility, which are also low in mass, the apparent resolving power is limited by contributions from ion gating. In contrast, the arrival-time distributions of low-mobility, native-like ions are not well explained using only contributions from ion gating and diffusion. For those species, the widths of arrival-time distributions are most consistent with the presence of multiple structures in the gas phase. [ABSTRACT FROM AUTHOR]

Iron oxide-based nanoparticles (NP) were covalently modified with sinapic acid (SA) through a condensation reaction to assist the ionization of both large and small molecules. The morphology of SA-modified NPs (SA-NP) was characterized by transmission electron microscopy (TEM), and the modification of the NP surface with SA was confirmed using ultraviolet (UV) and infrared (IR) spectroscopy. The number of SA molecules was estimated to be 6 per NP. SA-NP-assisted laser desorption/ionization was carried out on small molecules, such as pesticides and plant hormones, and large molecules, such as peptides and proteins. A peptide fragment from degraded proteins was detected more efficiently compared with conventional methods. [ABSTRACT FROM AUTHOR]

Site-specific protein modification is a widely-used biochemical tool. However, there are many challenges associated with the development of protein modification techniques, in particular, achieving site-specificity, reaction efficiency and versatility. The engineering of peptide ligases and their substrates has been used to address these challenges. This review will focus on sortase, peptidyl asparaginyl ligases (PALs) and variants of subtilisin; detailing how their inherent specificity has been utilised for site-specific protein modification. The review will explore how the engineering of these enzymes and substrates has led to increased reaction efficiency mainly due to enhanced catalytic activity and reduction of reversibility. It will also describe how engineering peptide ligases to broaden their substrate scope is opening up new opportunities to expand the biochemical toolkit, particularly through the development of techniques to conjugate multiple substrates site-specifically onto a protein using orthogonal peptide ligases. [ABSTRACT FROM AUTHOR]

Current materials research is inspired from biomineralization processes, where proteins control the formation of inorganic materials under soft reaction conditions. The implementation of biomineralization principles is a promising approach to generate tailored material properties for developing applications and devices. The results show the direct link between inorganic-binding peptides, biomineralization and the generation of new materials properties. Peptides which specifically bind to yttria-stabilized zirconia are selected by phage display and analyzed with regard to their amino acid composition. Based on these results the peptide 6-3-06 (SWNNMGWLPTFG) is selected for the peptide-assisted synthesis, generating biomineralized zirconia-based (BZO) material with photoluminescent properties. The material characterization by means of X-ray photoelectron spectroscopy reveals the incorporation of the peptide in the inorganic phase. More important this specific structure gives rise to photoluminescence (PL) in the blue wavelength region (~2.7 keV) at room temperature which is mainly attributed to states in the band gap introduced by organic–inorganic interfaces. This PL property is exclusive to the BZO whereas samples mineralized in the absence of the peptide did not show PL. The data clearly demonstrate that by biomineralization new material properties for technical materials can be generated. Importantly, this approach can be generalized for other inorganic materials. [ABSTRACT FROM AUTHOR]

Ultra-high pressure liquid chromatography (UHPLC) systems combined with state-of-the-art mass spectrometers have pushed the limit of deep proteome sequencing to new heights making it possible to identify thousands of proteins in a single LC-MS experiment within a few hours. The recently released EASY-spray technology allows one to implement nano-UHPLC with straightforwardness. In this work we initially characterized the EASY-spray containing a 50 cm column containing <2 μm particles and found that the system allowed 3000 proteins to be identified in 90 minutes. We then asked the question whether a fast and sensitive online 2D SCX-RP UHPLC-MS/MS workflow could compete with 1D long gradient analyses, using total analysis time versus proteome coverage and sample usage as benchmark parameters. The 2D LC-MS strategy consisted of the EASY-spray system that had been augmented by the addition of an SCX column. The conversion was made facile since no additional valves were required and by the use of components containing viper fittings. We benchmarked the system using a human cell lysate digest (<10 μg). The 2D SCX-RP UHPLC-MS/MS workflow allowed the identification of almost 37 000 unique peptides and 6000 proteins in a total analysis time of ∼7 hours. On the same system a 1D RP UHPLC-MS/MS workflow plateaued at only 20 000 peptides and 4400 unique proteins and required approx. 8 hours of analysis time. Furthermore, the 2D workflow could continue to increase the proteome coverage with longer analysis times, in fact with a 21 hour analysis we identified 56 600 unique peptides and >7500 proteins. We report, here, that with this fast online SCX-RP UHPLC-MS/MS workflow, the proteome coverage can be substantially extended without significantly compromising the analysis time and sample usage. [ABSTRACT FROM AUTHOR]

Plants produce a variety of proteins and peptides which are involved in their defense against pathogens. Serine protease inhibitors are a well-established class of inhibitors correlated with plant defense. Increased levels of protease inhibitors delay cell damage and expand the cell's life-span. Recently, the rapid emergence of antibiotic-resistant microbial pathogens has prompted immense interest in purifying novel antimicrobial proteins or peptides from plant sources. Usually, the purification of protease inhibitors is accomplished by salt-extraction, ultrafiltration and affinity chromatography. Here, we developed a novel approach based on iron oxide nanoparticles conjugated to dextran functionalized with trypsin beads that accelerate the quick screening and purification of antimicrobial peptides with serine protease inhibitor activity. The method described here also works for screening other inhibitors using particular protein kinases, and it is therefore a novel tool for use as the leading method in the development of novel antimicrobial agents with protease inhibitory activity. Finally, and no less important, molecular modelling and dynamics studies of a homologous inhibitor studied here with Escherichia coli trypsin and chymotrypsin are provided in order to shed some light on inhibitor–enzyme interactions. [ABSTRACT FROM AUTHOR]

O-Linked N-acetylglucosamine (O-GlcNAc) is an emerging post-translational modification (PTM) of proteins. Analysis of O-GlcNAc modification using mass spectrometry (MS) is often problematic because of the low stoichiometry of the modification. In this study, we developed a new method for enriching O-GlcNAc-modified peptides using reversible hydrazide chemistry. O-GlcNAc-modified peptides were first labeled with N-azidoacetylgalactosamine (GalNAz) using gatactosyltransferase-T1 (Y289L) enzyme. The azide group on the GalNAz residue was then reacted with 3-ethynylbenzaldehyde via copper-catalyzed Huisgen 1,3-cycloaddition “click reaction” to form an aromatic aldehyde group of glycopeptides. Aromatic aldehyde-derivatized glycopeptides were enriched by reversible hydrazone formation with hydrazide resin. Reaction conditions for each step, especially for the click reaction, were optimized to achieve complete reaction without significant side reactions. This method was validated using a tryptic digest of bovine α-crystallin, which is an O-GlcNAc-modified glycoprotein. The developed method was also applied to structure-specific enrichment of N-linked glycopeptides having non-reducing terminal GlcNAc residues. All materials and chemicals required for this method are commercially available and there is no need to prepare any special reagents, facilitating the introduction of this method in any laboratory. [ABSTRACT FROM AUTHOR]

Reflecting the increasing interest in combinatorial approaches, peptide phage display has seen an unprecedented expansion in a wide range of research areas. Its application to the discovery and analysis of metal binding peptides has opened up new research directions and largely contributed to the nanotechnology field. The rationale behind the need to identify such peptides varies depending on the final aim of the research and its application. Therefore, the possibility to modify the selection technique according to the different requirements would allow for a more systematic approach to be adopted and would ultimately provide substantial benefits. Although the standard panning method can be virtually applied to any target, its use for the identification of metal binding peptides does not provide the characteristics and the flexibility required for an efficient and tailored selection. Here we report on the development of a new panning method that can contribute to a faster, versatile and more informative analysis. Through the use of rolling-circle amplification, polymerase reaction and wild type phage, we have converted the standard selection technique into a more dynamic process in which adjustments can be evaluated and made consistently with the need of the experiment. The successfulness of the improved method is demonstrated in a number of panning experiments with different inorganic targets. The modifications applied to each selection are described and comparisons between the results obtained are made in order to extensively assess and evaluate the impact of the new process. The importance of tailoring the screening method to the specific objectives of a study is also considered. New binder sequences for the materials included in the investigation are identified; their sequences and distinctive characteristics are reported and their ability to act as templates for the nucleation of inorganic material is demonstrated and discussed. [ABSTRACT FROM AUTHOR]

Although halogen atoms can greatly improve the stability, selectivity, and bioactivity of proteins, direct halogenation of proteins or peptides by chemical strategy has been never achieved. Herein, we describe the developments of direct photochemical bromination and iodization of unprotected proteins and peptides in the direct irradiation device and the single-pulsed irradiation capillary reactor with biocompatible aqueous halides Br− and I−, respectively. These novel photochemical modifications are triggered by 193 nm laser photoexcitation of commonly photo-inert halide ions to form active radical species. High protein modification efficiency (>90%) can be achieved under just 10 ns ultra-short irradiation of a single pulse of laser shot while the compact native protein structure could be largely retained. The specifically modified residues are Tyr, His, Trp for bromination and Tyr, His for iodization. The photochemical halogenation sites and rates are highly selective to protein native structures, providing dynamic insights into protein structure alterations and protein–drug interfaces in human serum protein (HSA)–warfarin interaction. This novel 193 nm photochemical strategy opens new opportunities for the protein structure–function explorations. [ABSTRACT FROM AUTHOR]

Several methods can be used to improve the enrichment of phosphorylated proteins. In this paper, phosphopeptides were enriched using magnetic iron(ii,iii) oxide (magnetite, Fe3O4) nanoparticles (NPs) on a radiate microstructure silicon chip and then analyzed using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOFMS) without further purification processes. We have developed a radiate microstructure chip on which samples can be concentrated for analysis by MALDI-TOFMS. The phosphoprotein digests and magnetic iron oxide NPs aqueous solution were deposited onto the central zone of the radiate microstructure silicon chip and enabled the on-chip enrichment of phosphopeptides. Microscopic analysis confirmed that the applied samples were confined to the central zone. Sample spots focused on the chip were much smaller than those on an unmodified plate with the same total volume. Different additives were used and optimized processes were performed to minimize non-phosphopeptides interference. These data collectively demonstrate that our on-chip phosphopeptide enrichment protocol is a rapid and easy-to-use method for phosphoproteome analysis. [ABSTRACT FROM AUTHOR]

Self-assembled peptide and protein amyloid nanostructures have traditionally been considered only as pathological aggregates implicated in human neurodegenerative diseases. In more recent times, these nanostructures have found interesting applications as advanced materials in biomedicine, tissue engineering, renewable energy, environmental science, nanotechnology and material science, to name only a few fields. In all these applications, the final function depends on: (i) the specific mechanisms of protein aggregation, (ii) the hierarchical structure of the protein and peptide amyloids from the atomistic to mesoscopic length scales and (iii) the physical properties of the amyloids in the context of their surrounding environment (biological or artificial). In this review, we will discuss recent progress made in the field of functional and artificial amyloids and highlight connections between protein/peptide folding, unfolding and aggregation mechanisms, with the resulting amyloid structure and functionality. We also highlight current advances in the design and synthesis of amyloid-based biological and functional materials and identify new potential fields in which amyloid-based structures promise new breakthroughs. [ABSTRACT FROM AUTHOR]

One of the most attractive areas in inorganic chemistry is the synthesis of polyoxometalates (POMs) exhibiting new properties and applications. Since the impact of POMs in biochemistry and related fields of research has increased in the last few years, there has been a special interest in this topic. Significant progress in biological applications has been made where the interaction of POMs with amino acids, peptides and proteins is relevant. Versatile POMs play a series of different roles in the interaction with these biomolecules as described in this review. Various types of interactions are established, depending on the POM shape and charge, the amino acid side chain, peptide sequence or protein structure. Experimental conditions such as temperature, acidity, solvent, etc. are also important factors that influence the binding/reactivity of POM with biomolecules, as described herein. This understanding allows the adequate design of the POM-biomolecule couple for tailoring and controlling mechanisms of action such as catalysis, inhibition, and aggregation, or the crystallising agent. [ABSTRACT FROM AUTHOR]

A method for quantification of a pharmaceutical peptide in human plasma was developed using gradient elution LC-ICP-MS. A membrane desolvation (MD) system was applied to remove organic solvents from the eluent prior to the detection as SO+ in the dynamic reaction cell (DRC) of the ICP-DRC-MS instrument and subsequent quantification by post-column isotope dilution (IDA). Plasma proteins were precipitated prior to analysis. Analytical figures of merit including linearity, precision, LOD, LOQ and accuracy were considered satisfactory for analysis of plasma samples. The selectivity of the developed method was demonstrated for five pharmaceutically relevant peptides: desmopressin, penetratin, substance P, PTH (1-34) and insulin. Preliminary experiments on an ICP-MS/MS system using oxygen to reduce the effect of organic solvents were also performed to compare sensitivity. The results of the study demonstrated that LC-ICP-MS post-column IDA may constitute a valuable additional tool in quantification of non-labelled peptides in the early drug development offering absolute quantification without need of species specific standards. [ABSTRACT FROM AUTHOR]

A 12-mer peptide with an affinity for the meso diad sequence of poly(N-isopropylacrylamide) (PNIPAM) was identified through affinity-based peptide screening. A model protein (i.e., human serum albumin (HSA)) chemically modified with the peptide was successfully precipitated with PNIPAM above the lower critical solution temperature (LCST) of PNIPAM. [ABSTRACT FROM AUTHOR]

Conformational preferences of the 2-methylproline (2-MePro) residue and its role in stabilizing β-turn and polyproline II (PII) structures of peptides have been studied using density functional methods in the gas phase and in water. The population of the C7 H-bonded structures of Ac-2-MePro-NHMe is calculated to be dominant in the gas phase, whereas they become significantly depopulated and the populations of polyproline and α-helical structures increase in water. Due to the increased stability of the up-puckered PII structure, the populations of its PII and PI structures are more increased by 10.9% and decreased by 12.2% than Ac-Pro-NHMe in water, respectively. There is a large decrease in the cis population by 20.6% for the prolyl peptide bond by replacing Pro with 2-MePro in water, which is consistent with 13C NMR experiments. For Ac-Ala-2-MePro-NHMe, the conformer with a βI-turn is found to be the most preferred conformer with the population of 25.2% in water, whereas the open conformers are dominantly populated for Ac-Ala-Pro-NHMe. This is consistent with experimental results that the Ala-Pro sequence of the antigen mimotope associated with autoimmune recurrent thrombosis has been known to adopt a stable βI-turn structure with the replacement of Pro by 2-MePro in water. With the replacement of a single Pro by 2-MePro in the middle of the sequences of the polyproline model peptide Ac-(Pro)5-NMe2 and the single-strand model peptide of collagen Ac-(Hyp-Gly-Pro)2-NMe2, the relative stabilities of the PII structures are enhanced by 8.11 and 3.91 kcal mol−1, respectively, although there are only small changes in backbone torsion angles of both PII and PI structures. It can be deduced that the single strands of (Hyp-Gly-2-MePro)n with the stable PII structure might provide more stability to their triple helix. [ABSTRACT FROM AUTHOR]

We explore the use of magnetic relaxation sensors as an in vitroassay platform. Functionalized magnetic nanoparticles that exhibit transverse relaxation time (T2) changes with the introduction of an analyte have previously been used to detect proteins, antibodies, receptor ligands, peptides, nucleic acids, oligonucleotides, and other small molecules. The standard procedure for sensitizing particles towards specific targets is, however, time-consuming and cumbersome. We report here a new approach that exploits primary–secondary antibody binding for easily derivatizing particles against specific targets. The assay is shown to be quantitative and a multiplexed assay against three target analytes is demonstrated. [ABSTRACT FROM AUTHOR]

In nanopore analytics, individual molecules pass through a single nanopore giving rise to detectable temporary blockades in ionic pore current. Reflecting its simplicity, nanopore analytics has gained popularity and can be conducted with natural protein as well as man-made polymeric and inorganic pores. The spectrum of detectable analytes ranges from nucleic acids, peptides, proteins, and biomolecular complexes to organic polymers and small molecules. Apart from being an analytical tool, nanopores have developed into a general platform technology to investigate the biophysics, physicochemistry, and chemistry of individual molecules (critical review, 310 references). [ABSTRACT FROM AUTHOR]

The role of CuII-complexation of amyloidogenic peptides on the formation and structure of aggregates were analyzed in order to obtain a general insight into modulation of peptide aggregation by metal ions. The results mimicked various facets of biological relevant peptides/protein-aggregation. Important mechanistic features are the ability of CuII to associate the peptides by bridging coordination, the structural changes induced by CuII binding and their effect on peptide–peptide interactions. [ABSTRACT FROM AUTHOR]

A combination of two model approaches was used to explore the nature of silicone–protein interactions. Phage display technology was used to present a combinatorial library of peptides, in this case the phage library PhD.-12, to a series of model silicone surfaces. Solid, molecular cage silsesquioxanes were used as the models for silicone surfaces. The silsesquioxanes were octamethyloctasilsesquioxane (methyl8-T8), octaphenyloctasilsesquioxane (phenyl8-T8), octahydridooctasilsesquioxane (H8-T8) and dodecatrifluoropropyldodecasilsesquioxane (trifluoropropyl12-T12). The first two silsesquioxanes bear simple aliphatic (Me) and aromatic (Ph) pendant groups, and are simple silicone analogues. The second two silsesquioxanes have functionalised pendant groups, H and CF3CH2CH2. The panning results, using a wild-type phage as a control, show that the phage library binds to the simple aliphatic and aromatic silsesquioxanes strongly but non-specifically, and largely through the protein coat on the phage. The functionalised silsesquioxanes (H and CF3CH2CH2) are bound specifically by the phages. A statistical analysis of the DNA sequences of the strongly binding phages was carried out. The peptides that bind to H8-T8 are strongly enriched in proline content at positions 7 and 8, with enhanced histidine at positions 5 and 9, and enhanced threonine at position 11. The proline residues presumably induce a favourable conformation in the peptide for binding to the silsesquioxane surface. The enhanced histidine content is significant. It has been known for many years that imidazole has a particular affinity for electrophilic silanes. The amino acid distribution for trifluoropropyl12-T12 binding peptides is markedly different from that of H8-T8 silsesquioxane. Proline is again enhanced, but in positions 4, 11 and 12, and there are also very high concentrations of serine in positions 1, 9 and 11, and threonine in positions 1, 2 and 12. The highly polar nature of the CF3 group is likely to engage in hydrogen bonding with the OH groups of the serine and threonine side chains, accounting for the tight binding to trifluoropropyl12-T12 silsesquioxane. [ABSTRACT FROM AUTHOR]

A novel strategy to stabilize the α-helical secondary structures of peptides upon binding to gold nanoparticles is described. Using a model protein–protein interaction system, we showed that AuNPs decorated with stabilized p53 α-helix peptides can mediate specific molecular recognition with their target protein. [ABSTRACT FROM AUTHOR]

A label free fluorescence method for rapid detection of the presence of phosphorylated peptides/proteins in sample solutions using functional magnetic nanoparticles (MNPs) as the detection probes is demonstrated. [ABSTRACT FROM AUTHOR]

Homo- and heterochiral tetrameric γ-peptide derivatives in which the backbone is constrained by a five-membered ring populate a bend-ribbon conformation in solution stabilized by intramolecular hydrogen bonds. [ABSTRACT FROM AUTHOR]