An experimental and theoretical study of the gas-phase decomposition of monoprotonated peptide nucleic acids

Peptide nucleic acids (PNAs) are DNA/RNA mimics which have recently generated considerable interest due to their potential use as antisense and antigene therapeutics and as diagnostic and molecular biology tools. These synthetic biomolecules were designed with improved properties over corresponding oligonucleotides such as greater binding affinity to complementary nucleic acids, enhanced cellular uptake, and greater stability in biological systems. Because of the stability and unique structure of PNAs, traditional sequence confirmation methods are not effective. Alternatively, electrospray ionization coupled with Fourier transform ion cyclotron resonance mass spectrometry shows great potential as a tool for the characterization and structural elucidation of these oligonucleotide analogs. Extensive gas-phase fragmentation studies of a mixed nucleobase 4-mer (AACT) and a mixed nucleobase 4-mer with an acetylated N-terminus (N-acetylated AACT) have been performed. Gas-phase collision-induced dissociation of PNAs resulted in water loss, cleavage of the methylene carbonyl linker containing a nucleobase, cleavage of the peptide bond, and the loss of nucleobases. These studies show that the fragmentation behavior of PNAs resembles that of both peptides and oligonucleotides. Molecular mechanics (MM+), semiempirical (AM1), and ab initio (STO-3G) calculations were used to investigate the site of protonation and determine potential low energy conformations. Computational methods were also employed to study prospective intramolecular interactions and provide insight into potential fragmentation mechanisms.     

Theoretical studies on the vibrational spectra, thermodynamic properties, detonation properties, and pyrolysis mechanisms for polynitroadamantanes

To look for high energy density materials (HEDM), the relationships between the structures and the performances of polynitroadamantanes (PNAs) were studied. The assigned infrared spectra of PNAs obtained at the density functional theory (DFT) B3LYP/6-31G level were used to compute the thermodynamic properties on the basis of the principle of statistical thermodynamics. The thermodynamic properties are linearly related with the number of nitro groups as well as with the temperatures. Detonation properties of PNAs were evaluated by using the Kamlet-Jacobs equation based on the calculated densities and heats of formation for titled compounds, and it is found that only when the number of nitro groups of PNA is equal to or more than eight can it be possible for PNAs to be used as HEDMs. The relative stabilities of PNAs were studied by the pyrolysis mechanism using the UHF-PM3 method. The homolysis of the C-NO2 bond is predicted to be the initial step of thermal decomposition. The activation energies (Ea) for the homolysis decrease with the number of nitro groups being increased on the whole. The stability order of dinitroadamantane isomers derived from the interactions among nitro groups is consistent with what is determined by Ea. The relations between the Ea's and the electronic structure parameters were discussed. In combination with the stability, PNA (1,2,3,4,5,6,7,8,9,10-) is recommended as the target of HEDM with insensitivity.     

PNA: Synthetic Polyamide Nucleic Acids with Unusual Binding Properties

The astonishing discovery that peptide nucleic acids (PNAs, B=nucleobase), in spite of their drastic structural difference to natural DNA, are better nucleic acid mimetics than many other oligonucleotides has resulted in an explosion of research into this class of compounds. The synthesis, physical properties, and biological interactions of PNAs as well as their chimeras with DNA and RNA are summarized here.     

Nucleopeptide Assemblies Selectively Sequester ATP in Cancer Cells to Increase the Efficacy of Doxorubicin

Herein, we report that assemblies of nucleopeptides selectively sequester ATP in complex conditions (for example, serum and cytosol). We developed assemblies of nucleopeptides that selectively sequester ATP over ADP. Counteracting enzymes interconvert ATP and ADP to modulate the nanostructures formed by the nucleopeptides and the nucleotides. The nucleopeptides, sequestering ATP effectively in cells, slow down efflux pumps in multidrug‐resistant cancer cells, thus boosting the efficacy of doxorubicin, an anticancer drug. Investigation of 11 nucleopeptides (including d ‐ and l ‐enantiomers) yields five more nucleopeptides that differentiate ATP and ADP through either precipitation or gelation. As the first example of assemblies of nucleopeptides that interact with ATP and disrupt intracellular ATP dynamics, this work illustrates the use of supramolecular assemblies to interact with small and essential biological molecules for controlling cell behavior.     

Synthesis of nucleopeptides by employing an enzyme-labile urethane protecting group

Nucleoproteins are naturally occurring biopolymers in which the hydroxy group of a serine, a threonine, or a tyrosine moiety is linked through a phosphodiester group to the 3'- or 5'-end of a nucleic acid. For the study of the biological phenomena in which nucleoproteins are involved, for example, viral replication, nucleopeptides embodying the characteristic linkage between the peptide chain and the oligonucleotide may serve as powerful tools. However, as a result of the multifunctionality and the pronounced acid and base lability of nucleopeptides, their synthesis requires the application of a variety of orthogonally stable blocking groups, which can be removed under the mildest conditions. We have developed a new mild enzymatic deprotection method, that is, the penicillin G acylase-catalyzed hydrolysis of the N-phenylacetoxybenzyloxycarbony (PhAcOZ) group, for the synthesis of nucleopeptides. We demonstrate the wide applicability of this method by coupling the N-terminally deprotected nucleopeptides 31 a-c with PhAcOZ-protected amino acids and subsequent removal of the N-PhAcOZ group from fully protected nucleotetrapeptides 32 a,b with penicillin G acylase. The reaction conditions are very mild (pH 6.8) so that no undesired side reaction such as cleavage of the nucleotide bond or beta-elimination of the nucleotide was observed.     

Impact of C-Terminal Chemistry on Self-Assembled Morphology of Guanosine Containing Nucleopeptides

Herein, we report the design and characterization of guanosine-containing self-assembling nucleopeptides that form nanosheets and nanofibers. Through spectroscopy and microscopy analysis, we propose that the peptide component of the nucleopeptide drives the assembly into β-sheet structures with hydrogen-bonded guanosine forming additional secondary structures cooperatively within the peptide framework. Interestingly, the distinct supramolecular morphologies are driven not by metal cation responsiveness common to guanine-based materials, but by the C-terminal peptide chemistry. This work highlights the structural diversity of self-assembling nucleopeptides and will help advance the development of applications for these supramolecular guanosine-containing nucleopeptides.     

Food analysis and food authentication by peptide nucleic acid (PNA)-based technologies

This tutorial review will address the issue of DNA determination in food by using Peptide Nucleic Acid (PNA) probes with different technological platforms, with a particular emphasis on the applications devoted to food authentication. After an introduction aimed at describing PNAs structure, binding properties and their use as genetic probes, the review will then focus specifically on the use of PNAs in the field of food analysis. In particular, the following issues will be considered: detection of genetically modified organisms (GMOs), of hidden allergens, of microbial pathogens and determination of ingredient authenticity. Finally, the future perspectives for the use of PNAs in food analysis will be briefly discussed according to the most recent developments.     

Charge dependent behavior of PNA/DNA/PNA triplexes in the gas phase

Intact noncovalent complexes were studied in the gas phase using negative ion nano-ESI mass spectrometry. Among various noncovalent systems studied in the gas phase, the interaction of DNA strands with peptide nucleic acids (PNAs) presents a strong interest as biologically relevant systems. PNAs originally described by Nielsen are used as DNA mimics as possible medical agents by imprisoning DNA single strands into stable noncovalent complexes. Two types of PNAs were investigated in the PNA/DNA multiplex: the original Nielsen's PNA and a modified backbone PNA by the introduction of syn- and anti-(aminoethyl)thiazolidine rings. We first investigated the stoichiometry of PNA/DNA multiplexes formed in solution and observed them in the gas phase via qualitative kinetics of complementary strand associations. It resulted in observing PNA2/DNA triplexes (ts) as the multiply deprotonated species, most stable in both the solution and gas phase. Second, charge-dependant decompositions of these species were undertaken under low-energy collision conditions. It appears that covalent bond cleavages (base releasing or skeleton cleavage) occur from lower ts charge states rather than ts unzipping, which takes place from higher charge states. This behavior can be explained by considering the presence of zwitterions depending on the charge state. They result in strong salt-bridge interactions between the positively charged PNA side chain and the negatively charged DNA backbone. We propose a general model to clearly display the involved patterns in the noncovalent triplex decompositions. Third, the relative stability of three PNA2/DNA complexes was scrutinized in the gas phase by acquiring the breakdown curves of their ts(6-) form, corresponding to the ts unzipping. The chemical structures of the studied PNAs were chosen in order to evidence the possible influence of backbone stereochemistry on the rigidity of PNA2/DNA complexes. It provided significantly different stabilities via V(m) measurements. The relative gas-phase stability order obtained was compared to that found in solution by Chassaing et al., and shows qualitative agreement.     

DNA and RNA binding properties of an arginine-based ‘Extended Chiral Box’ Peptide Nucleic Acid

Lys-based ‘chiral box’ Peptide Nucleic Acids (PNAs with three adjacent 2D-Lys-based chiral monomers) have shown unsurpassed specificity in DNA recognition. In this Letter, the binding performances of arginine-based chiral PNAs were evaluated for PNAs containing in the middle part of the strand either a 2D,5L-Arg monomer or three adjacent 2D-; 2D,5L-; 5L-Arg monomers (‘Extended Chiral Box’), a combination never studied before. The binding performances of the PNAs were studied by evaluating the melting temperatures of fullmatch and mismatch PNA-DNA and PNA-RNA hybrids and by studying their structure by circular dichroism (CD). The data indicated that the arginine side chains inserted in the PNA structure are perfectly equivalent to lysine side chains as far as oligonucleotide recognition is concerned. The insertion of an ‘Extended Chiral Box’ into PNA differently influences the binding properties to DNA and RNA: the additional side chains had no observable effect on binding affinity and selectivity toward DNA, whereas, seemed to slightly disturb the binding affinity to RNA but at the same time highly enhancing the recognition selectivity.     

Mixing Biomimetic Heterodimers of Nucleopeptides to Generate Biocompatible and Biostable Supramolecular Hydrogels

As a new class of biomaterials, most supramolecular hydrogels formed by small peptides require the attachment of long alkyl chains, multiple aromatic groups, or strong electrostatic interactions. Based on the fact that the most abundant protein assemblies in nature are dimeric, we select short peptide sequences from the interface of a heterodimer of proteins with known crystal structure to conjugate with nucleobases to form nucleopeptides. Being driven mainly by hydrogen bonds, the nucleopeptides self‐assemble to form nanofibers, which results in supramolecular hydrogels upon simple mixing of two distinct nucleopeptides in water. Moreover, besides being biocompatible to mammalian cells, the heterodimer of the nucleopeptides exhibit excellent proteolytic resistance against proteinase K. This work illustrates a new and rational approach to create soft biomaterials by a supramolecular hydrogelation triggered by mixing heterodimeric nucleopeptides.     

Charge transfer through modified peptide nucleic acids

We studied the charge transfer properties of bipyridine-modified peptide nucleic acid (PNA) in the absence and presence of Zn(II). Characterization of the PNA in solution showed that Zn(II) interacts with the bipyridine ligands, but the stability of the duplexes was not affected significantly by the binding of Zn(II). The charge transfer properties of these molecules were examined by electrochemistry for self-assembled monolayers of ferrocene-terminated PNAs and by conductive probe atomic force microscopy for cysteine-terminated PNAs. Both electrochemical and single molecular studies showed that the bipyridine modification and Zn(II) binding do not affect significantly the charge transfer of the PNA duplexes.     

The alpha-helical peptide nucleic acid concept: merger of peptide secondary structure and codified nucleic acid recognition

A novel platform for nucleic acid recognition that integrates the alpha-helix secondary structure of peptides with the codified base-pairing capability of nucleic acids is reported. The resulting alpha-helical peptide nucleic acids (alpha PNAs) are composed of a repeating tetrapeptidyl unit, aa(1)-aa(2)-aa(3)-Ser(B), where aa(1) through aa(3) represent generic ancillary amino acids and B = nucleobases linked to Ser via a methylene bridge. Effective syntheses of constituent Fmoc-protected nucleoamino acids (Fmoc-Ser(B)-OH, where B = thymine, cytosine, and uracil) are described along with a protocol for the solid-phase synthesis of 21mer alpha PNAs containing five such nucleobases. By varying the ancillary amino acids, two distinct classes of alpha PNAs were constructed, having a net charge of -1 or +6, respectively, at physiological pH. The modular nature of the alpha PNA platform was illustrated by the synthesis of symmetrical disulfide-bridged alpha PNA dimers containing 10 nucleobases. Hybridization of these alpha PNAs with ssDNA has been examined by thermal denaturation, gel electrophoresis, and circular dichroism (CD) and the data indicated that alpha PNA binds to ssDNA in a cooperative manner with high affinity and sequence specificity. In general, b2 alpha PNAs bind faster and more strongly with ssDNA than do the corresponding b1 alpha PNAs. Parallel alpha PNA-DNA complexes are more stable than their antiparallel counterparts. CD studies also revealed that the hybridization event involves the folding of both species into their helical conformations. Finally, NMR experiments provided conclusive evidence of Watson-Crick base pairing in alpha PNA-ssDNA hybrids.     

Rapid Synthesis of Propargyl-γ-Modified Peptide Nucleic Acid Monomers for Late-Stage Functionalization of Oligomers

The exceptional hybridization properties of peptide nucleic acids (PNAs) coupled with the ease of their synthesis has made this artificial nucleic acid mimetic a desirable platform for diagnostics, therapeutics and supramolecular engineering. PNA backbone modifications have been extensively explored to finetune physicochemical properties and for conjugation of functional molecules. Here, we detail the synthesis of a universal γ-propargyl-PNA backbone from serine, and its acylation with the four DNA canonical nucleobases. The availability of serine as d or l enantiomer provide simple accesses to PNA oligomers for hybridization with natural oligonucleotides or for orthogonal hybridization circuitry. We show that late-stage conjugation enables optimization of the physicochemical properties. This approach is appealing due to its orthogonality to Fmoc-SPPS, its flexibility and ease for introducing diversity by on-resin copper(I)-catalyzed azide-alkyne cycloaddition (CuAAC). We exemplified the utility of these novel monomers with PNA based hybridization chain reactions (HCRs).     

酞菁一富勒烯给受体体系研究进展

酞菁的大π共轭体系使其具有良好的电子给予能力和对可见光的强烈吸收性质,可作为人工光合作用体系中的光捕捉单元．富勒烯具有独特的三维笼状结构,几乎所有反应中都具有低的重组能,是良好的电子或能量受体．将富勒烯与酞菁结合起来,集两个光活性实体于一身,由于酞菁和富勒烯给受体之间的电子相互作用,又赋予其新的性质,具有良好的应用前景,是富勒烯和酞菁领域中的研究热点．本文将酞菁．富勒烯给受体体系分为共价和非共价两种连接方式,介绍其结构与性质以及两者的相互关系,总结了其在有机太阳能电池、非线性光学和液晶性等方面的应用进展,展望了该体系的未来发展趋势．     

Metal-containing peptide nucleic acid conjugates

Peptide Nucleic Acids (PNAs) are non-natural DNA/RNA analogues with favourable physico-chemical properties and promising applications. Discovered nearly 20 years ago, PNAs have recently re-gained quite a lot of attention. In this Perspective article, we discuss the latest advances on the preparation and utilisation of PNA monomers and oligomers containing metal complexes. These metal- conjugates have found applications in various research fields such as in the sequence-specific detection of nucleic acids, in the hydrolysis of nucleic acids and peptides, as radioactive probes or as modulators of PNA·DNA hybrid stability, and last but not least as probes for molecular and cell biology.     

PNA-DNA duplexes, triplexes, and quadruplexes are stabilized with trans-cyclopentane units

Peptide nucleic acids (PNAs) are non-natural nucleic acid mimics that bind to complementary DNA and RNA with high affinity and selectivity. PNA can bind to nucleic acids in a number of different ways. Currently, the formation of PNA-oligonucleotide duplex, triplex, and quadruplex structures have been reported. PNAs have been used in numerous biomedicial applications, but there are few strategies to predictably improve the binding properties of PNAs by backbone modification. We have been studying the benefits of incorporating (S,S)-trans-cyclopentane diamine units (tcyp) into the PNA backbone. In this Communication, we report the improvement in stability associated with tcyp incorporation into PNA-DNA duplexes, triplexes, and quadruplexes. The broad utility of this modification across multiple types of PNA structures is unique and should prove useful in the development of applications that rely on PNA.     

Caprin-1 Promotes Cellular Uptake of Nucleic Acids with Backbone and Sequence Discrimination

The cellular delivery of oligonucleotides has been a major obstacle in the development of therapeutic antisense agents. PNAs (Peptide Nucleic Acid) are unique in providing a modular peptidic backbone that is amenable to structural and charge modulation. While cationic PNAs have been shown to be taken up by cells more efficiently than neutral PNAs, the generality of uptake across different nucleobase sequences has never been tested. Herein, we quantified the relative uptake of PNAs across a library of 10 000 sequences for two different PNA backbones (cationic and neutral) and identified sequences with high uptake and low uptake. We used the high uptake sequence as a bait for target identification, leading to the discovery that a protein, caprin-1, binds to PNA with backbone and sequence discrimination. We further showed that purified caprin-1 added to cell cultures enhanced the cellular uptake of PNA as well as DNA and RNA.     

Nanocrystals modified with peptide nucleic acids (PNAs) for selective self-assembly and DNA detection

Gold nanocrystals modified with peptide nucleic acids (PNAs) have been prepared and applied to self-assembly and DNA sensing. Experiments with different PNA structural motifs show that (1). the versatility in PNA synthetic design can be used to modulate the electrostatic surface properties of nanocrystals, presenting an opportunity to control assembly rate and aggregate size, (2). short (6 base) PNAs can hybridize effectively while attached to nanoparticles, providing a route to generating materials with small interparticle spacings, and (3). the superior base pair mismatch selectivity of PNAs is further enhanced on nanosurfaces, enabling PNA-modified nanoparticles to act as highly selective nanoscale sensors, as well as synthons for defect-free self-assembly. This last feature was coupled with a substantial change in colloidal stability upon DNA hybridization to develop a novel colorimetric DNA assay that detects the presence of single base imperfections within minutes. Various modes of PNA hybridization, including the first practical application of PNA-PNA interactions, were used to direct the assembly of nanoparticles into macroscopic arrangements. Shorter duplex interconnects and greater specificity in assembly were obtained compared to similar experiments with DNA-modified nanocrystals.     

(S,S)-trans-cyclopentane-constrained peptide nucleic acids. a general backbone modification that improves binding affinity and sequence specificity

Replacing the ethylenediamine portion of aminoethylglycine peptide nucleic acids (aegPNAs) with one or more (S,S)-trans-cyclopentane diamine units significantly increases binding affinity and sequence specificity to complementary DNA, making these modified PNAs ideal for use as nucleic acid probes in genomic analysis. The synthesis and study of this new class of PNAs (tcypPNAs) is described in which trans-cyclopentane diamine has been incorporated into several positions, and in varying number, within PNA backbones of mixed-base sequences.     

Peptides of DL-willardiine. homo- and heterodipeptides from willardiine

One of the types of pyurinylpeptides (i.e., peptides constructed of pyrimidyl-and purinyl--amino acids) consists of peptides including residues of only those -amino acids the lateral radicals of which contain nucleic bases. Below, such amino acids will be called nucleoamino acids and the peptides derived from them nucleopeptides. The latter are hybrid analogs of nucleic acids and proteins. Their study may lead to important theoretical and practical results.Translated from Khimiya Geterotsiklicheskikh Soedinenii, Vol. 6, No. 7,pp. 1002–1003, July, 1970.We are continuing the synthesis and study of the properties of the pyurinylpeptides and nucleopeptides.