KR-20260065658-A - Aptamer specifically binding to Patulin and use thereof

Abstract

The present invention relates to an aptamer that specifically binds to patulin and its uses. As a result of efforts to develop an aptamer with high binding affinity to patulin, an aptamer with excellent specificity and binding affinity even to small target molecules was discovered in silico, which significantly reduces time and cost compared to conventional typical SELEX. Furthermore, it was confirmed that the discovered aptamer exhibits excellent high sensitivity and specificity for patulin, so the aptamer of the present invention can selectively detect only the target patulin in a sample containing a minute amount of patulin.

Inventors

• 윤순일
• 찬드란 크리스나라지
• 아스마리 미스가나 맹기스투

Assignees

• 전북대학교산학협력단

Dates

Publication Date

20260511

Application Date

20241101

Claims (6)

1. An aptamer that specifically binds to patulin, comprising the nucleotide sequence of SEQ ID NO. 1.
2. A composition for detecting patulin comprising the aptamer of claim 1.
3. A kit for detecting patulin comprising the aptamer of claim 1.
4. A sensor for detecting patulin comprising the aptamer of claim 1 and a substrate on which the aptamer is immobilized.
5. A method for detecting patulin comprising the step of treating a sample with the aptamer of claim 1.
6. A method for detecting patulin according to claim 5, wherein the sample is taken from water, soil, waste, food, the inside of an animal’s intestine, or the tissue of an animal or plant.

Description

Aptamer specifically binding to Patulin and use thereof The present invention relates to an aptamer that specifically binds to patulin and its uses. Mycotoxins are secondary metabolites produced by molds that, even in trace amounts through contaminated agricultural products and food, cause direct harm to humans and livestock as toxic, carcinogenic, and mutagenic agents. Furthermore, some toxins remain in the tissues of livestock products or milk, and are substances with a high potential to cause secondary harm to humans. These mycotoxins are produced by molds during the growth, storage, and distribution of agricultural products. Mycotoxin production is influenced by environmental factors such as temperature and humidity, and once produced, the toxins are heat-stable and do not decompose even after cooking or processing. Tens of thousands of mold species exist in nature, producing over 300 types of mycotoxins; among these, approximately 10 to 20 types can contaminate food and animal feed, potentially causing harmful effects on human health. Mycotoxin contamination in agricultural products cannot be completely avoided, and according to the FAO, more than 25% of the world's crops are reported to be infected with mycotoxins. In this regard, the WHO and others assess that the risk posed by these mycotoxins is actually higher than that of food additives or pesticide residues among food-related hazards (Mycotoxins in food, Ed. N. Magan and M. Olsen, Woodhead Publishing Ltd and CRC Press LLC (2004)), and internationally, more than 99 countries regulate about a dozen types, including aflatoxins, patulin, ochratoxin A (OTA), deoxynivalenol, fumonisines, zearalenone, T-2 toxin, sterigmatocystin, HT-2 toxin, diacetoxyscirpenol, ergot alkaloids, phomopsins, and agaric acid; currently, Korea also regulates aflatoxins, patulin, Fumonisin, ochratoxin, deoxynivalenol, and zealarenone are regulated in agri-food and animal feed. Meanwhile, as awareness of the importance of patulin has grown, various studies are being conducted to prevent resulting health problems and crop losses. One such study concerns the development of aptamers that specifically bind to patulin to safely remove or inactivate it. Aptamers are small oligonucleotides that bind specifically and selectively to target molecules. As is widely known, nucleic acids are linear polymers in which nucleotides are linked by covalent bonds; generally, nucleic acids such as DNA and RNA serve as storage vessels for information to express proteins with cellular structural and enzymatic activities. Meanwhile, nucleic acids form three-dimensional structures and become stabilized by forming complexes through interactions with specific substances; due to this characteristic, nucleic acids can act as ligands for specific substances, including proteins. Accordingly, aptamers that bind to specific substances with high binding affinity and specificity can be produced from a library of single-stranded nucleic acids arranged in various base sequences through a specific selection process and base sequence determination. Furthermore, unlike antibodies, which are conventional sensing materials used in the sensor field, aptamers are nucleic acid structures, so they exhibit excellent thermal stability. Since they are synthesized in vitro, they do not require animals or cells, making them economical in terms of production costs. Additionally, because there are no restrictions on target substances, aptamers can be synthesized for a wide range of targets, from biomolecules such as proteins and amino acids to low-molecular-weight organic chemicals like endocrine disruptors, antibiotics, and pharmaceutical residues, as well as bacteria and viruses. Therefore, aptamers for various target substances are being produced, and due to the characteristics of aptamers that bind to target substances with specificity and strong affinity, much research has recently been conducted on applying aptamers to new drug development, drug delivery systems, and biosensors. In addition, aptamers are highly suitable materials for introduction into detection methods for trace amounts of residual pesticides, and they can also be applied to detect specific residual pesticide substances through nanobiotechnology utilizing them. Figure 1 is the secondary structure of a patulin aptamer. Figure 2 shows the interactions of the patulin-aptamer complex after docking, where π-π interactions are indicated by green dashes and hydrogen bond interactions are indicated by blue lines. Figure 3 shows the RMSD plot of the patulin aptamer structural deviation based on the initial form. Figure 4 shows the hydrogen bonds formed between patulin and aptamer during a 100ns MD simulation. Figures 5 and 6 show the results of electrochemical characteristics confirming the patulin detection ability of the DNA-aptamer of the present invention. (5a) shows the cyclic voltammetry (CVs) recorded at a scan rate of 50 mV s⁻¹ . (5b) shows the electroche