肽核酸—基因调控的利器

肽核酸(PNA)是具有类多肽骨架的DNA类似物,PNA的主链骨架是由N(2-氨基乙基)-甘氨酸与核酸碱基通过亚甲基羰基连接而成的。PNA可以特异性地与DNA或RNA杂交,形成稳定的复合体。PNA由于其自身的特点可以对DNA复制、基因转录、翻译等进行有针对的调控,同时作为杂交探针大大提高了遗传学检测和医疗诊断的效率和灵敏度。肽核酸（PNA）特异性地识别和结合互补核酸序列被引进用于医学和生物学的研究,展示了其独特的生化属性,成为了基因奥秘的探索者。     

Preparation and evaluation of a peptide nucleic acid gene chip system associated with surface plasmon resonance

The aim of this study was to construct a gene chip system based on a surface plasmon resonance technique, where peptide nucleic acid (PNA) oligomers are used as probes. Since the self-assembled monolayer (SAM) technology offers good control at the molecular level, we prepared 2D surface chemistry via SAM for probe attachments. PNA, which was designed according to the bioinformatics, was immobilized on the SAM-modified chip, and subsequently, relevant parameters of the experiment were ensured and optimized. Our results suggest that the ion strength and pH value of the buffer solution do not play significant roles in PNA or its complementary strand hybridization. The PNA probe binds to its complementary nucleic acid strand with a higher sensitivity and specificity compared to those of a traditional DNA probe. The PNA probe combined with surface plasmon resonance (SPR) technology has the benefits of being a label-free and in-real time monitor, as well as having improved hybridization and stability efficiency, which highlight the PNA gene chip detection system as a promising biosensor for clinical applications.     

肽核酸在分子生物学技术中的应用

肽核酸(PNA)作为一种人工合成的核酸类似物,以中性的肽链酰胺2-氨基乙基甘氨酸键取代了DNA中的戊糖磷酸二酯键骨架,其余部分与DNA相同。PNA可通过Watson-Crick碱基配对的形式识别并结合DNA或RNA序列,形成稳定的双螺旋结构。与传统的DNA或RNA相比,PNA具有生物学稳定性高、杂交特异性强、杂合体的稳定性高和杂交速度快等明显优点,使PNA具有良好的物理化学性质和生物学特性,在检测目的核酸序列中单碱基突变、PCR基因分子诊断与检测、荧光原位杂交定量分析、基因芯片和生物传感器技术等调控水平和临床应用上有自己的特点。简要综述了近年来肽核酸在上述分子生物学技术中的运用以及应用前景的展望。     

修饰性肽核酸的细胞转运

肽核酸是人工合成的寡核苷酸类似物,以N-(2-氨乙基)甘氨酸结构单元替代DNA分子中的戊糖-磷酸结构。与天然核酸相比,肽核酸可以更高效地与DNA或RNA特异性杂交,在分子生物学和基因药物领域具有良好的应用前景。但是,肽核酸骨架呈电中性,难以高效穿过细胞膜,这成为工程应用的最大障碍。为了改善肽核酸的细胞转运性能,对肽核酸进行化学修饰是近年来的研究热点。结合近十年来文献报道和本实验室的工作,对肽核酸的骨架修饰和配合物结合修饰两类增强细胞转运的修饰方法进行综述,并对修饰性肽核酸细胞转运研究中存在的问题以及未来的研究趋势及其应用提出了见解。     

Modulation of DNA and RNA by PNA

Peptide nucleic acid (PNA), a synthetic DNA mimic that is devoid of the (deoxy)ribose-phosphate backbone yet still perfectly retains the ability to recognize natural nucleic acids in a sequence-specific fashion, can be employed as a tool to modulate gene expressions via several different mechanisms. The unique strength of PNA compared to other oligonucleotide analogs is its ability to bind to nucleic acid targets with secondary structures such as double-stranded and quadruplex DNA as well as RNA. This digest aims to introduce general readers to the advancement in the area of modulation of DNA/RNA functions by PNA, its current status and future research opportunities, with emphasis on recent progress in new targeting modes of structured DNA/RNA by PNA and PNA-mediated gene editing.     

Antisense properties of peptide nucleic acid

Peptide nucleic acid (PNA) is a nucleic acid mimic in which the deoxyribose phosphate backbone has been replaced by a pseudo-peptide polymer to which the nucleobases are linked. PNA-oligomers can be synthesized in relatively large amounts, are highly stable in biological environments, and bind complementary DNA and RNA targets with remarkably high affinity and specificity. Thus PNA possesses many of the properties desired for a good antisense agent. Until recently, limited uptake of PNA into cells has been the major obstacle for applying PNA as an antisense agent in cell cultures and in vivo. Here, the antisense properties of PNA in vitro and in vivo will be reviewed. In particular, we will focus on recent observations indicating that PNA equipped with or without various uptake moieties may function as an efficient and gene-specific inhibitor of translation in Escherichia coli and in certain mammalian cell types.     

Unwinding of unnatural substrates by a DNA helicase

Helicases separate double-stranded DNA into single-stranded DNA intermediates that are required during replication and recombination. These enzymes are believed to transduce free energy available from ATPase activity to unwind the duplex and translocate along the nucleic acid lattice. The nature of enzyme-substrate interactions between helicases and duplex DNA substrates has not been well-defined. Most helicases require a single-stranded DNA overhang adjacent to duplex DNA in order to initiate unwinding. The strand containing the overhang is referred to as the loading strand whereas the complementary strand is referred to as the displaced strand. We have investigated the interactions between a DNA helicase and the DNA substrate by replacing the displaced strand with a nucleic acid mimic, peptide nucleic acid (PNA). PNA is capable of forming duplex structures with DNA according to Watson-Crick base pairing rules, but contains a N-(2-aminoethyl)glycine backbone in place of the deoxyribose phosphates. The PNA-DNA hybrids had higher melting temperatures than their DNA-DNA counterparts. Dda helicase, from bacteriophage T4, was able to unwind the DNA-PNA substrates at similar rates as DNA-DNA substrates. The results indicate that the rate-limiting step for unwinding is relatively insensitive to the chemical nature of the displaced strand and the thermal stability of oligonucleotide substrates.     

肽核酸(peptide nucleic acid,PNA)阵列

肽核酸(PNA)以N—(2—氨基乙基)甘氨酸替代DNA分子中的磷酸戊糖骨架。它能特异性地识别与DNA、RNA所形成的杂交体。PNA—DNA、PNA—RNA的热稳定性要比相应的DNA—DNA、DNA—RNA高,而且PNA识别单碱基的能力强于DNA和RNA,使之在微阵列,尤其是SNP检测领域有着广泛的应用前景。本文简述了PNA阵列从探针设计、阵列合成、杂交和检测的全过程。     

Peptide nucleic acid-anthraquinone conjugates: strand invasion and photoinduced cleavage of duplex DNA.

A bis-peptide nucleic acid (PNA)-anthraquinone imide (AQI) conjugate has been synthesized and shown to form strand invasion complexes with a duplex DNA target. The two arms of the bis-PNA each consist of five consecutive thymine residues and are linked by a flexible, hydrophilic spacer. Probing with potassium permanganate reveals that the bis-PNA complexes to duplex DNA at A5.T5sites with local displacement of the T5DNA strand. The 5 bp sequence targeted by the PNA is the shortest strand invasion complex reported to date. Irradiation of the strand invasion complex results in asymmetric cleavage of the displaced strand, with more efficient cleavage at the 3'-end of the loop. This result indicates that the bis-PNA binds to the DNA such that the C-terminal T5sequence forms the strand invasion complex, leaving the N-terminal T5sequence to bind by triplex formation, thereby placing the AQI closer to the 3'-end of the displaced strand, consistent with the observed photocleavage pattern. The ability of the PNA to directly report its binding site by photoinduced cleavage could have significant utility in mapping the secondary and tertiary structure of nucleic acids.     

Peptide nucleic acids: deoxyribonucleic acid mimics with a peptide backbone

This tutorial briefly describes a new class of synthetic biopolymer, which is referred to as peptide nucleic acid (PNA). In PNA, individual nucleobases are linked to an achiral neutral peptide backbone. PNA exhibits the hybridization characteristic (e.g., Watson—Crick duplex formation) of DNA. The achiral peptide backbone provides similar interbase distances as natural DNA, and adequate flexibility to permit base pair interactions with complementary RNA or DNA strands. Several potential applications of PNA oligomers in biotechnology are suggested. These include the use of PNAs as a probe for specific recognition of a DNA or RNA sequence selective, purification of nucleic acids via designed high affinity binding to PNA, screening for DNA mutations, and as possible therapeutic agents.     

Recognition of Double-Stranded Dna by Peptide Nucleic Acid

Abstract

Peptide nucleic acid (PNA) is an oligonucleotide mimic in which the backbone of DNA has been replaced by a pseudopeptide. We here show that there are distinct variations as to how PNA oligomers interact with double-stranded DNA depending on choice of nucleobases. Thymine-rich homopyrimidine PNA oligomers recognise double-stranded polynucleotides by forming PNA₂-DNA triplexes with the DNA purine strand. By contrast, cytosine-rich homopyrimidine PNAs add to double-stranded polynucleotides as Hoogsteen strands, forming PNA-DNA₂ triplexes, while homopurine, or alternating thymine-guanine, PNA oligomers invade DNA to form PNA-DNA duplexes.     

γPNA一种新型高效的肽核酸

肽核酸(peptide nucleic acid,PNA)是一种人工合成的具有类多肽骨架的DNA类似物,具有与核酸结合特异性强、组织和细胞内生物稳定性好、半衰期长等优点。通过靶向结合DNA/RNA而抑制其复制、转录和翻译过程,进行基因调控。在PNA骨架结构中γ位点引入带手性的官能团,能形成右手螺旋结构,显著提高其与靶DNA/RNA的杂交特性,这种PNA衍生物称之为γPNA。γPNA的溶解性、热稳定性和特异性等化学与生物学特性明显改善,在基因编辑和作为探针检测等方面具有良好的应用前景。通过对γPNA结构、性质及其研究进展进行总结,以期为γPNA反义应用提供理论依据和参考。     

肽核酸——一种新型的反义物质

肽核酸是以肽为骨架的一种新型ＤＮＡ模拟物。已经证明肽核酸具有与ＤＮＡ和ＲＮＡ结合的高度亲合性、良好的稳定性及能方便地固相合成等特性。在反义技术和基因治疗中有着很好的前景。本文综述了肽核酸的生物化学特性及其在反义技术方面的应用。     

Unique properties of purine/pyrimidine asymmetric PNA.DNA duplexes: differential stabilization of PNA.DNA duplexes by purines in the PNA strand

PNA.DNA duplexes are significantly stabilized by purine nucleobases in the PNA strand. To elucidate and understand the effect of switching the backbone in a nucleic acid duplex, we now report a thermodynamics study along with a solution conformations study of two purine/pyrimidine strand asymmetric duplexes and a strand symmetrical control by comparing the behavior of all four possible PNA/DNA combinations. In essence, we are comparing an identical basepair stack connected by either an aminoethyl glycine PNA or a deoxyribose DNA backbone. We show that the PNA.DNA duplexes containing purine-rich PNA strands are stabilized with regard to the thermal melting temperature and free energy as well as enthalpy (and concomitantly relatively less entropically disfavored). Based on our data, we find it unlikely that differences in counterion binding (identical ionic-strength dependence was observed), hydration (identical and insignificant water release was observed), or single-strand conformation can be responsible for the difference in duplex stability. The only consistent difference observed between the purine-rich PNA versus the pyrimidine-rich PNA in isosequential PNA.DNA duplexes is the significant increase in both binding enthalpy and entropy for the PNA.DNA duplexes containing pyrimidine-rich PNA in organic solvent, which would indicate that these duplexes are relatively enthalpically disfavored in water. Although our results so far do not allow us to identify the origin of the different stabilities of homopurine/homopyrimidine PNA.DNA duplexes, the evidence does point to a significant structural component, which involves enthalpic contributions both within the duplex structure and also from bound water molecules.