Transmission of extramembrane conformational change into current: construction of metal-gated ion channel

Interaction with Fe(III) induces the reversible conformational switch of the extramembrane segment in the artificial receptor channel, which is transmitted into membranes as an increase in channel current (ion flux).     

Modulating ion channel properties of transmembrane peptide nanotubes through heteromeric supramolecular assemblies

A selective heteromeric supramolecular assembly process is devised to create functional single channels of altered ion conductance, charge selectivity, and rectification. The hollow transmembrane tubular structure produced spontaneously from the self-assembly of cyclic-d,l-alpha-peptides in lipid bilayers is modified by designer cyclic peptide "cap" subunits that bind site-selectively at the mouth of the channel assembly.     

Controlling the activity of peptides and proteins with smart nucleic acid-protein hybrids

Oligonucleotide-peptide conjugates have frequently been used to control the localisation of the conjugate molecule. For example, the oligonucleotide segment has allowed spatially addressed immobilization of peptides and proteins on DNA-arrays via hybridisation while the peptide part has most frequently been used to confer transfer of oligonucleotide cargo into live cells. The regulation of functional properties such as the affinity of these bioconjugates for protein targets has rarely been addressed. This review article describes the current developments in the application of smart oligonucleotide-peptide hybrids. The mutual recognition between nucleic acid segments is used to constrain the structure or control the distance between peptide and protein segments. Application of these new type of oligonucleotide-peptide hybrids allowed not only the regulation of binding affinity of peptide ligands but also control of enzymatic and optical activity of proteins.     

Thermosensitive diblock copolymers with designed molecular weight distribution: Synthesis by continuous living cationic polymerization and micellization behavior

The synthesis of diblock copolymers with designed molecular weight distributions (MWDs) was successfully demonstrated in a continuous living cationic polymerization system using simple equipment. The control of MWDs was achieved by gradually feeding a polymerization reaction mixture into a terminating agent. As thermosensitive diblock copolymers, poly(vinyl ethers) containing a thermosensitive segment with oxyethylene side chains and a hydrophilic segment were prepared. The polymerization was carried out in a gas‐tight microsyringe, and the polymerization mixture was added continuously into methanol during the second‐stage polymerization. The self‐association behavior of the resulting diblock copolymers was evaluated by dynamic light scattering in water. MWD‐designed polymers with thermosensitive segments that varied continuously in length and hydrophilic segments of nearly uniform lengths formed micelles with a broad size distribution. Conversely, polymers with nearly uniform thermosensitive segments and hydrophilic segments of different lengths formed micelles with a narrow size distribution, as observed with conventional narrow MWD diblock copolymers. Thus, the MWD of the thermosensitive segment proved a decisive factor in achieving fine control of self‐association. © 2008 Wiley Periodicals, Inc. J Polym Sci Part A: Polym Chem 46: 2212–2221, 2008     

Tandem ligation of unprotected peptides through thiaprolyl and cysteinyl bonds in water.

Tandem ligation for the synthesis and modification of proteins entails forming two or more regiospecific amide bonds of multiple free peptide segments without a protecting-group scheme. We here describe a semi-orthogonal strategy for ligating three unprotected peptide segments, two of which contain N-terminal (NT) cysteine, to form in tandem two amide bonds, an Xaa-SPro (thiaproline), and then an Xaa-Cys. This strategy exploits the strong preference of an NT-cysteinyl peptide under acidic conditions to undergo selectively an SPro-imine ligation rather than a Cys-thioester ligation. Operationally, it was performed in the N --> C direction, first by an imine ligation at pH < 3 to afford an Xaa-thiazolidine ester bond between a peptide containing a carboxyl terminal (CT)-glycoaldehyde ester and a second peptide containing both an NT-Cys and a CT-thioester. The newly created O-ester-linked segment with a CT-thioester was then ligated to another NT-cysteinyl peptide through thioester ligation at pH > 7 to form an Xaa-Cys bond. Concurrently, this basic condition also catalyzed the O,N-acyl migration of an Xaa-thiazolidine ester to the Xaa-SPro bond at the first ligation site to complete the tandem three-segment ligation. Both ligation reactions were performed in aqueous buffered solvents. The effectiveness of this three-segment ligation strategy was tested in six peptides ranging from 19 to 70 amino acids, including thiaproline --> proline analogues of somatostatins and two CC-chemokines. The thiaproline replacements in these peptides and proteins did not result in altered biological activity. By eliminating the protecting-group scheme and coupling reagents, tandem ligation of multiple free peptide segments in aqueous solutions enhances the scope of protein synthesis and may provide a useful approach for combinatorial segment synthesis.     

Continuously Tunable Ion Rectification and Conductance in Submicrochannels Stemming from Thermoresponsive Polymer Self‐Assembly

A biomimetic conical submicrochannel (tip side ca. 400 nm) with functions of continuously tunable ion rectification and conductance based on thermoresponsive polymer layer‐by‐layer (LbL) self‐assembly is presented. These self‐assembled polymers with different layers exhibited a capability to regulate the effective channel diameter, and different ion rectifications/conductance were achieved. By controlling temperature, the conformation and wettability of the assembled polymers were reversibly transformed, thus the ion rectification/conductance could be further adjusted subtly. Owing to the synergistic effect, the ion conductance could be tuned over a wide range spanning three orders of magnitude. Moreover, the proposed system can be applied for on‐demand on‐off molecule delivery, which was important for disease therapy. This study opens a new door for regulating channel size according to actual demand and sensing big targets with different size with one channel.     

Structural characterization of sialic acid synthase by electrospray mass spectrometry—A tetrameric enzyme composed of dimeric dimers

Sialic acid synthase (NeuB) encoded by the neuB gene catalyzes the condensation of N-acetylmannosamine and phospho(enol)pyruvate to form N-acetylneuraminic acid. The enzyme is essential for the biosynthesis of polysialic acid, a capsular sugar polymer functioning as a virulent factor and antiphagocytic barrier. This report demonstrates the first characterization on the quaternary structure of NeuB from Escherichia coli (EcNeuB) and Streptococcus agalactiae (SaNeuB) by nanoflow electrospray ionization mass spectrometry (ESI-MS). Under non-denaturing conditions, Tris buffer was observed to induce a higher ratio of tetramer/dimer of NeuB in the ESI mass spectra, providing supportive evidence for the existence of a "structurally-specific" tetramer. The instrument parameters were found to significantly affect the ratio of detected tetramer/dimer in ESI mass spectra. The harshest conditions, including high desolvation voltages and pressure in the collision cell, led to enhanced detection of the 160 kDa tetramer. The prevalence of dimeric form is likely the cause in loss of tetramer stability in gas-phase arising from insufficient collisional cooling, which implies an asymmetric assembly, possibly composed of dimeric dimers. Most interestingly, the hypothesis was further supported by chemical cross-linking of SaNeuB, in which the reaction of shorter linker yielded mainly the dimer whereas that of longer linker produced both dimer and tetramer. Furthermore, the ESI-MS analysis can reflect dramatic change of pH-dependent quaternary structure in association with enzyme activity, suggesting the tetrameric form may be the primary species responsible for the enzyme catalysis.     

Studies on peptides. CLXVII. Solid-phase syntheses and immunological properties of fragment peptides related to human malaria circumsporozoite protein

A glycine-linked tetramer of Asn-Ala-Asn-Pro, a tandem repeated sequence of malaria circumsporozoite (CS) protein, was synthesized by the Boc-based solid phase method, followed by deprotection with 1 M trimethylsilyl trifluoromethanesulfonate-thioanisole in trifluoroacetic acid. In addition, three tetramer-related peptides were similarly synthesized, i.e., a 34-residue peptide [linked with TH, a proposed T-cell epitope of CS, at the C-terminus of the tetramer], a 46-residue peptide and a 59-residue peptide [linked with HA or HA', two proposed T-cell epitopes of influenza hemagglutinin protein, at the N-terminus of the above 34-residue peptide]. Their immunological properties were examined by enzyme-linked immunosorbent assay, for which three different congenic strains of mouse were used to raise the specific antibodies. Despite conjugation of T-cell epitopes to the tetramer, the mice of low-responder strains to the tetramer failed to produce any antibody specific to the tetramer. However, with the aid of recombinant interleukin 2 as an adjuvant, the low-responder mice produced antibody with relatively high titers.     

Transformation of peptide nanotubes into a vesicle via fusion driven by stereo-complex formation

Two types of peptide nanotubes, one is prepared from an amphiphilic peptide having a right-handed helix segment and the other from that having a left-handed helix segment, are shown to transform the morphology into a vesicle by membrane fusion due to stereo-complex formation between these helical segments.     

Biomimetic and aggregation-driven crystallization route for room-temperature material synthesis: growth of beta-Ga(2)O(3) nanoparticles on peptide assemblies as nanoreactors

The room-temperature synthesis of beta-Ga2O3 nanocrystal was examined by coupling two biomimetic crystallization techniques, enzymatic peptide nanoassembly templating and aggregation-driven crystallization. The catalytic template of peptide assembly nucleated and mineralized primary beta-Ga2O3 crystals and then fused them to grow single-crystalline and monodisperse nanoparticles in the cavity of the peptide assembly at room temperature. In this work, the peptide assembly was exploited as a nanoreactor with an enzymatic functionality catalyzing the hydrolysis of gallium precursors. In addition, the characteristic ring structure of peptide assembly is expected to provide an efficient dehydration pathway and crystallization control over the surface tension, which are advantageous for beta-Ga2O3 crystal growth. This multifunctional peptide assembly could be applied for syntheses of a variety of nanomaterials that are kinetically difficult to grow at room temperature.     

Molecular design and synthesis of artificial ion channels based on cyclic peptides containing unnatural amino acids

A series of novel cyclic peptides composed of 3 to 5 dipeptide units with alternating natural-unnatural amino acid units, have been designed and synthesized, employing 5-(N-alkanoylamino)-3-aminobenzoic acid with a long alkanoyl chain as the unnatural amino acid. All cyclic peptides with systematically varying pore size, shape, and lipophilicity are found to form ion channels with a conductance of ca. 9 pS in aqueous KCl (500 mM) upon examination by the voltage clamp method. These peptide channels are cation selective with the permeability ratio P(Cl(-))/P(K(+)) of around 0.17. The ion channels formed by the neutral, cationic, and anionic cyclic peptides containing L-alanine, L-lysine, and L-aspartate, respectively, show the monovalent cation selectivity with the permeability ratio P(Na(+))/P(K(+)) of ca. 0.39. On the basis of structural information provided by voltage-dependent blockade of the single channel current of all the tested peptides by Ca(2+), we inferred that each channel is formed from a dimer of the peptide with its peptide ring constructing the channel entrance and its alkanoyl chains lining across the membrane to build up the channel pore. The experimental results are consistent with an idea that the rate of ion conduction is determined by the nature of the hydrophobic alkanoyl chain region, which is common to all the channels.     

Total Synthesis of Siomycin A: Completion of the Total Synthesis

The total synthesis of siomycin A ( 1 ), a representative compound of the thiostrepton family of peptide antibiotics, was achieved by incorporating the five synthetic segments A ( 2 ), B ( 3 ), C ( 4 ), D ( 5 ), and E ( 6 ). The dehydropiperidine segment A ( 2 ) was esterified with the dihydroquinoline segment C ( 4 ), and the subsequent coupling with the β‐phenylselenoalanine dipeptide segment D ( 5 ) at the segment C portion followed by lactamization between the segments A and D gave segment A‐C‐D ( 27 ). This was amidated with the pentapeptide segment B ( 3 ) at the segment A portion followed by one‐pot cyclization (between segments A and B) and elongation (with the β‐phenylselenoalanine dipeptide segment E ( 6 ) at the segment A portion), thus furnishing siomycin A ( 1 ).     

Multicomponent supramolecular thermoplastic elastomer with peptide‐modified nanofibers

Bioactive nanofibers present a promising synthetic niche for in vivo applications due to their morphological and functional resemblance to the extracellular matrix. Potentially interesting nanofibers are constructed from the hard‐segment regimes in well‐defined thermoplastic elastomers (TPEs). The supramolecular interactions between these hard segments cause physical crosslinking by the formation of nanofibers and provide excellent mechanical properties. Here, we make use of a new class of biocompatible supramolecular TPEs, in which both the formation of the main chain and the hard block is based on multiple hydrogen‐bonding interactions. A self‐assembly process is explored to arrive at well‐defined peptide‐modified nanofibers embedded in a biocompatible soft matrix. Crucial for the success in the synthetic design is the use of an exact match between the molecular recognition units of the peptide and the supramolecular unit that takes care of forming the supramolecular nanofibers of the TPE. Evidence for the strong anchoring of the modified peptides in the hard‐segment nanofibers of the supramolecular TPE is provided by simple extraction experiments. © 2011 Wiley Periodicals, Inc. J Polym Sci Part A: Polym Chem, 2011     

Peptides derived from two dynamically disordered proteins self-assemble into amyloid-like fibrils

Short peptides derived from p14ARF and Hdm2 (14 and 15 amino acids in length, respectively), two cancer associated proteins, have been found to co-assemble into amyloid-like structures. Larger protein domains containing these peptide segments interact in cells and also undergo a disorder-to-order transition upon binding in vitro. In contrast to the association of beta-strand assemblies with amyloid diseases, the system described herein utilizes the formation of binary, extended beta-strands as a novel mechanism of biomolecular assembly. The beta-strand-containing fibrils formed from these peptides may allow the directed assembly of decorated fibrils with applications as biological nanostructures.     

Electrophoretic chip for fractionation of selective DNA fragment

A microchannel chip has been used to fractionate selected segments from an electrophoretic flow of separated fragments. A sample, which covers the size from 35 to 670 bp, was initially separated using an 8.8-cm-long channel at the electric field strength of 100 V/cm. The target fragment of 318 bp was selected and extracted from the separation channel. High-resolution fractionation was achieved by introducing new procedures for blocking, extraction, and segment transfer. Fractionation quality with and without blocking were compared using a 310 Genetic Analyzer (Applied Biosystems). The results show that no contamination was found in the sample, which was fractionated with blocking; however, a contamination by short segments was found in the sample, which was fractionated without blocking. Furthermore, fractionation by the chip was found to be of higher fidelity than that by the polyacrylamide slab gel, which displayed a small overlapped peak after the target peak. Compared with the traditional method, our chips enable faster and high-fidelity fractionation, thus providing a new tool for bioanalysis and other applications.     

Identification of Core Segment of Amyloidal Peptide Mediated by Chaperone Molecules by using Scanning Tunneling Microscopy

We illustrate in this work that pristine assemblies of amyloidal peptides can be obtained by perturbations of reduced scanning bias, and show a broad distribution in peptide length. In contrast, the chaperone‐mediated peptide co‐assembly presents ordered lamellar structures with a homogeneous distribution in length, which could be attributed to the core segment of the peptide. The efforts are beneficial for gaining insight into the aggregation propensity of peptides and inter‐peptide interactions.     

Electrophoretic chip for high-fidelity fractionation of double-stranded DNA

We report the high fidelity, on-chip fractionation of selected segments from an electrophoretic flow of separated fragments. dsDNA fragments (10-330 base pairs (bp)) were initially separated using a 6.5 cm long channel with an electric field strength of 150 V/cm. As an example of the fractionation process, a target fragment of 20 bp was selected and extracted from the separation channel. The extraction was confirmed and evaluated by fluorescence imaging. High resolution and extraction fidelity were achieved by introducing new procedures for (i) extraction channel-blocking and (ii) segment transfer with cleaning. These procedures are necessary for the development of a practical, fully automated multitarget fractionation electrophoretic chip. A kind of CCD image processing method was introduced to monitor, control, and evaluate the procedure of fractionation. The resolution limits of the separation and extraction are discussed.     

α‐Peptide–Oligourea Chimeras: Stabilization of Short α‐Helices by Non‐Peptide Helical Foldamers

Short α‐peptides with less than 10 residues generally display a low propensity to nucleate stable helical conformations. While various strategies to stabilize peptide helices have been previously reported, the ability of non‐peptide helical foldamers to stabilize α‐helices when fused to short α‐peptide segments has not been investigated. Towards this end, structural investigations into a series of chimeric oligomers obtained by joining aliphatic oligoureas to the C‐ or N‐termini of α‐peptides are described. All chimeras were found to be fully helical, with as few as 2 (or 3) urea units sufficient to propagate an α‐helical conformation in the fused peptide segment. The remarkable compatibility of α‐peptides with oligoureas described here, along with the simplicity of the approach, highlights the potential of interfacing natural and non‐peptide backbones as a means to further control the behavior of α‐peptides.     

利用单片机直接驱动光柱显示器件

介绍一种单片机直接驱动光柱显示器件的方法.该方法采用动态扫描的形式直接驱动两个101线光柱.光柱的十位数叫做段码,个位数叫做线数.与段码对应的段叫做当前段,段码减一的段及其以下的段叫做十线全显段.关显示就是关双光柱.