Compound for recognizing notched G-quadruplex, and preparation method and application of compound

A technology for quadruplexes and compounds, which is applied in the field of compounds that identify gapped G-quadruplexes, can solve problems such as large differences, incomplete gapped G-quadruplex planes, and inability to recognize gapped G-quadruplexes, etc., to achieve The effect of increasing stability and enhancing the ability to inhibit DNA replication

Inactive Publication Date: 2020-08-11
SUN YAT SEN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The plane of the gapped G-quadruplex is incomplete, and its structure is quite different from the previously discovered regular G-quadruplex. Therefore, the previously discovered compounds that bind to the regular G-quadruplex cannot recognize the gapped G-quadruplex
Since the discovery of the gapped G-quadruplex, compounds that can effectively recognize it have not been reported

Method used

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  • Compound for recognizing notched G-quadruplex, and preparation method and application of compound
  • Compound for recognizing notched G-quadruplex, and preparation method and application of compound
  • Compound for recognizing notched G-quadruplex, and preparation method and application of compound

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0037] Combining guanine with peptides to prepare compounds that recognize gapped G-quadruplexes:

[0038] The raw material of guanine is peptide nucleic acid PNA-G, and the polypeptide that binds to the G-quadruplex plane is derived from human RHAU protein. The full sequence is HPGHLKGREIGMWYAKKQGQKNK (N-terminal to C-terminal) (SEQ ID NO.1), and the synthesis of polypeptide-modified guanine It is completed on a polypeptide synthesizer, and the specific steps of the synthesis are prior art, and will not be repeated here. The synthesis can be completed in any polypeptide synthesis company. The compound that recognizes the gap G-quadruplex provided in this example was completed in Beijing Saibaisheng Company. The final sequence is G-HPGHLKGREIGMWYAKKQGQKNK (SEQ ID NO.2) (wherein G is PNA- G, at the N-terminus of the polypeptide). For the convenience of description, the compound that recognizes the gap G-quadruplex prepared by the guanine and the polypeptide provided in this em...

Embodiment 2

[0040] Validation that gRPCs specifically recognize gapped G-quadruplexes:

[0041] In this example, EMSA experiment is used to verify that GRPC specifically recognizes the gap G-quadruplex. Among them, MYOG and ABTB2 were selected as gapped G-quadruplexes, both of which have three layers of guanine planes, with a gap on the 5' end and 3' end planes, respectively. In this example, the sequences of MYOG and ABTB2 are shown in Table 1, wherein MYOG 3332 and ABTB2 3233 are gapped G-quadruplexes, MYOG 3333 and ABTB2 3333 are complete three-layer G-quadruplexes after mutation, and MYOG 2332 and ABTB2 2233 are mutated complete two-layer G-quadruplexes. The 5' end of the DNA has the fluorescent group FAM.

[0042] Table 1

[0043]

[0044] In the present embodiment, the steps of EMSA experiment are as follows:

[0045] (1) 0.01 μM DNA was dissolved in buffer, and the buffer composition was 20 mM Tris-HCl buffer (pH7.4), 50 mM KCl and 40% (wt / vol) PEG 200.

[0046] (2) DNA sampl...

Embodiment 3

[0053] Verify that the guanine of GRPC fills the gap and forms a hydrogen bond:

[0054] After GRPC binds to the gap G-quadruplex, guanine in GRPC will fill in the gap and undergo hydrogen bond interaction. In this example, DMS protection experiments are used to verify that guanine in GRPC fills in the gap and forms hydrogen bonds.

[0055] The operation procedure of DMS protection experiment is as follows:

[0056] (1) 0.05 μM MYOG 3332 and ABTB2 3233 DNA were dissolved in 200 μL buffer, the buffer components were 50 mM Lithium cacodylate (pH 7.4), 40% (wt / vol) PEG 200, 50 mM LiCl or KCl.

[0057] (2) DNA samples were heated at 95°C for 5 minutes, and then cooled slowly to 20°C.

[0058] (3) After the sample was equilibrated at room temperature for 20 minutes, 4 μL of DMS was added, shaken and mixed, and left for 6 minutes.

[0059] (4) Add final concentrations of 100 μM mercaptoethanol, 0.3 M sodium acetate (pH 5.2), 10 ug salmon sperm DNA, and place on ice to terminate th...

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Abstract

The invention relates to the field of biology, and discloses a compound for recognizing a notched G-quadruplex, and a preparation method and application of the compound. The compound for recognizing the notched G-quadruplex comprises a guanine molecule and a polypeptide or a small molecule capable of recognizing the G-quadruplex, wherein the polypeptide or the small molecule is connected with theguanine molecule through an amido bond, and the structural formula of the compound is shown in the specification. In the structural formula, R1 or R2 is the small molecule or the polypeptide. According to the compound for recognizing the notched G-quadruplex provided by the invention, a notch is filled with guanine to form a hydrogen bond, and the polypeptide or the small molecule is combined witha guanine plane at the tail end of the notched G-quadruplex so as to allow two binding sites to exist between the polypeptide or the small molecule and the notched G-quadruplex, so the notched G-quadruplex can be specifically recognized, the stability of the notched G-quadruplex can be improved, and the DNA replication inhibition ability of the notched G-quadruplex can be enhanced.

Description

technical field [0001] The invention relates to the field of biology, in particular to a compound for recognizing gap G-quadruplex, its preparation method and application. Background technique [0002] Single-stranded DNA or RNA rich in guanine can form a structure called a G-quadruplex. Different from the common DNA double helix structure, the hydrogen bonds that maintain the G-quadruplex structure mainly occur between guanines, and the four guanines that undergo hydrogen bonding form a guanine plane. The G-quadruplex formation sequence in the human genome is mainly distributed in the telomere of the chromosome, the promoter of the gene, and the first intron. In RNA, the G-quadruplex forming sequence is concentrated in the 5'UTR and 3'UTR of mRNA, and exists in a variety of ncRNAs. [0003] The G-quadruplex is involved in DNA replication, gene transcription, mRNA translation and other processes. G-quadruplexes can also lead to genome instability in some G-quadruplex-rela...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N9/14C07D473/18
CPCC07D473/18C12N9/14
Inventor 郑克威何意得
Owner SUN YAT SEN UNIV
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