Method for controlling length of amyloid fibrils through pressurized cutting process

Abstract

The present invention relates to a method for controlling the length of amyloid fibrils through a pressurized cutting process. Specifically, according to the present invention, long amyloid fibrils having a relatively long length can be uniformly cut into a desired specific length, and thus fibrils having a predetermined length can be produced precisely and reproducibly, thereby enabling cytotoxicity analysis according to the length of amyloid fibrils in neurodegenerative disease research. In addition, amyloid fibrils according to the present invention consistently maintain electrical properties, mechanical properties, and chemical reactivity due to the uniform and consistent length thereof, and thus may be suitable for commercial applications. In addition, since the pressurized cutting process of the present invention is relatively simple and economical, the commercial potential of various products and services using amyloid fibrils can be increased. Therefore, the pressurized cutting process is highly likely to be used in various technical fields such as neurodegenerative disease research, biosensors, filtration systems, nano-electronic devices, nanoparticles for drug screening, and bioadhesives.

Description

Method for controlling amyloid fibrillar length through a pressure cutting process

[0001] The present invention relates to a method for controlling the length of amyloid fibrils through a pressure cutting process.

[0002] Amyloid fibrils are known to be one of the pathological causes of neurodegenerative diseases such as Alzheimer's disease, Parkinson's disease, and Huntington's disease. In these diseases, proteins aggregate to form amyloid fibers, which induce toxicity in cells and cause tissue damage (Fig. 1). Meanwhile, previous studies have primarily generated amyloid fibers through spontaneous or chemical induction, but these methods could not precisely control the length of the fibers. As a result, research on how the length of amyloid fibers affects disease progression has been limited, and it has been difficult to obtain a clear answer as to how fibers of a specific length induce cytotoxicity.

[0003] Due to their unique structural properties, amyloid fibers hold significant potential for application in the fields of nanotechnology and materials science. Amyloid fibers of uniform length can be utilized in nano-based technologies such as electronic fibers, biosensors, and filtration systems (Fig. 2). Furthermore, amyloid fibers possess a highly stable β-sheet structure, which prevents them from deforming in various chemical and physical environments. Thanks to these characteristics, amyloid fibers are considered ideal materials for various applications requiring precise nanoscale structures. However, their use in nanotechnology applications has been limited because the length of amyloid fibers cannot be precisely controlled.

[0004] Furthermore, amyloid fibers have faced difficulties in structural analysis studies due to their structural diversity (polymorphism). Amyloid fibers possess a structure consisting of repeating stacks of β-sheets and can form crystal-like arrangements under specific conditions. Consequently, while amyloid fibers can be studied precisely using tools such as cryo-EM, X-ray diffraction, and high-resolution NMR, such analysis has been challenging due to the fibers' non-uniform lengths. Additionally, in disease research, amyloid fibers of specific lengths can play a crucial role in toxicity mechanisms or inter-fiber interactions. However, conventional methods have struggled to generate fibers of uniform length, making it difficult to conduct sufficient structural analysis related to diseases.

[0005] Accordingly, the inventors sought to solve the above problem by developing a method to precisely control the length of amyloid fibers using a pressure cutting process method capable of adjusting filter size and the number of extrusions, while simultaneously enabling mass production of amyloid fibers cut to a predetermined length.

[0006] The present invention aims to provide a method for controlling the length of amyloid fibrils that not only enables obtaining relatively long amyloid fibers in a desired uniform short fiber form through the repetition of a pressure cutting process, but also allows for mass production using a simple process.

[0007] In addition, length-controlled amyloid fibers produced according to the above method can be widely utilized in various technological fields, such as research on neurodegenerative diseases, biosensors, filtration systems, nanoelectronic devices, nanoparticles for drug screening, and bioadhesives.

[0008] The technical problems that the present invention aims to solve are not limited to those mentioned above, and other unmentioned technical problems will be clearly understood by those skilled in the art to which the present invention belongs from the description below.

[0009] The present specification provides a method for controlling the length of amyloid fibrils, comprising: a) a step of synthesizing amyloid long fibers; and b) a step of extruding the synthesized amyloid long fibers and cutting them into amyloid short fibers of a predetermined length by passing them through a filter having a plurality of pores.

[0010] For example, the amyloid intestinal fiber may be formed by forming a fiber aggregate of one protein selected from egg white lysozyme (HEWL), Abeta42, Tau(AcPHF6), alpha-synuclein, insulin, and combinations thereof.

[0011] For example, step a above may be performed by heating a solution in which purified protein is dissolved to a temperature range of 60 to 90°C under conditions of a pH range of 2 to 7, and then maintaining it at a predetermined temperature to form a fiber shape.

[0012] For example, in step b above, the length of the short fibers can be controlled by controlling one or more of the number of extrusions, filter pore size, and amyloid long fiber concentration.

[0013] For example, in step b above, the filter may have one material selected from polycarbonate, anodic aluminum oxide, polyethersulfone, and combinations thereof.

[0014] For example, in step b above, the pores of the filter may have a diameter in the range of 100 to 400 nm.

[0015] For example, the extrusion of step b above can be performed at a pressure in the range of 100 to 500 psi and an extrusion speed in the range of 2 to 5 mL / min.

[0016] For example, the extrusion of step b above can be repeated 1 to 120 times.

[0017] For example, the concentration of amyloid fibers extruded in step b above may be in the range of 0.1 to 2 wt% based on the total weight of the solution containing amyloid fibers.

[0018] For example, the extrusion of amyloid long fibers in step b above can be performed under conditions maintained in a temperature range of 20 to 25 ℃.

[0019] For example, the amyloid short fibers prepared according to the pressure cutting method process including steps a and b above may have a loss rate of 20% or less based on the original fiber.

[0020] Additionally, the present specification provides length-controlled amyloid fibers prepared according to the above method, wherein the amyloid fibers have a standard deviation of length within the range of 18 to 23% based on atomic force microscopy (AFM) analysis of 300 or more individual amyloid fibers.

[0021] For example, the amyloid fibers mentioned above can be utilized in one or more technical fields selected from neurodegenerative disease research, biosensors, filtration systems, nanoelectronic devices, nanoparticles for drug screening, and bioadhesives.

[0022] According to the present invention, relatively long amyloid fibers can be uniformly cut to a specific desired length, thereby enabling the precise and reproducible production of fibers of a predetermined length, and consequently, it becomes possible to analyze cytotoxicity based on the length of amyloid fibers in the study of neurodegenerative diseases.

[0023] In addition, the amyloid fibers according to the present invention are suitable for commercial applications because their electrical properties, mechanical properties, and chemical reactivity are consistently maintained due to their uniform and constant length.

[0024] Furthermore, the amyloid fibers produced according to the present invention possess uniform length and structural consistency, which minimizes polymorphism—a variable in high-resolution structural analysis methods such as cryo-EM and X-ray diffraction analysis—and enables the acquisition of more precise data. Accordingly, it is possible to deepen the understanding of the amyloid fiber formation process, structural characteristics, and toxicity mechanisms.

[0025] Furthermore, the pressure cutting process of the present invention is relatively simple and economical, which can increase the commercialization potential of various products and services using amyloid fibers. Accordingly, it has great potential for application in various technological fields, such as research on neurodegenerative diseases, biosensors, filtration systems, nanoelectronic devices, nanoparticles for drug screening, and bioadhesives.

[0026] Figure 1 schematically shows the physiological structure of the brain and nerve cells of a normal or Alzheimer's disease patient.

[0027] FIG. 2 schematically illustrates the application fields in which the amyloid fiber of one embodiment of the present invention can be utilized, including (a) a heavy metal filter, (b) nanoparticles for amyloid oligomer drug screening, and (c) a bioadhesive.

[0028] FIG. 3 schematically illustrates (a) a pressure cutting process using a carbonate filter and an extruder, and (b) a pressure cutting mechanism according to an embodiment of the present invention.

[0029] FIG. 4 shows (a) a pressure cutting process cycle of amyloid fibers according to one embodiment of the present invention, (b) atomic force microscope (AFM) images of the cut amyloid fibers according to the number of cycles, (c) diameter and length according to the number of cycles, and (d) the results of statistical analysis thereof.

[0030] FIG. 5 shows (a) an AFM image according to filter pore size during a pressure cutting process according to an embodiment of the present invention, (b) and (c) the diameter and length of the fabricated amyloid fibers and the results of statistical analysis thereof.

[0031] FIG. 6 shows (a) an AFM image of amyloid adsorbed on a filter after a pressure cutting process according to an embodiment of the present invention, (b) and (c) the adsorption tendency of amyloid fibers according to the filter pore structure and a schematic diagram thereof, and (d) and (e) the protein quantification (BCA assay) results and loss rate of the fabricated 100 and 200 nm amyloid fibers.

[0032] Figure 7 shows the results of confirming changes in diameter and length of amyloid fibers according to an embodiment and a comparative example of the present invention after inducing binding and aggregation between amyloid fibers by reacting them at 24°C for 2 weeks.

[0033] FIG. 8 shows the lengths produced after varying the amyloid long fiber concentration during the pressure cutting process according to an embodiment of the present invention and the results of their statistical analysis.

[0034] Hereinafter, embodiments of the present invention are described in detail with reference to the attached drawings so that those skilled in the art can easily implement the present invention. However, the present invention may be embodied in various different forms and is not limited to the embodiments disclosed below. Furthermore, in order to clearly disclose the present invention in the drawings, parts unrelated to the present invention have been omitted, and identical or similar reference numerals in the drawings indicate identical or similar components.

[0035] The purpose and effects of the present invention may be naturally understood or become clearer from the following description, and the purpose and effects of the present invention are not limited to the description below alone.

[0036] The objectives, features, and advantages of the present invention will become clearer through the following detailed description. Furthermore, in describing the present invention, if it is determined that a detailed description of known technology related to the present invention may unnecessarily obscure the essence of the invention, such detailed description will be omitted. Hereinafter, embodiments according to the present invention will be described in detail with reference to the attached drawings.

[0037] Meanwhile, the terms "long fiber" and "short fiber" used throughout the specification and claims of the present invention should be understood as being for the purpose of comparing relative sizes.

[0038]

[0039] Amyloid fibrils are known to be a pathological cause of neurodegenerative diseases such as Alzheimer's disease, Parkinson's disease, and Huntington's disease. For the study of these diseases, amyloid fibers were previously generated through spontaneous or chemical induction; however, these methods made it difficult to precisely control fiber length, making it challenging to accurately determine how fiber length affects disease progression. Meanwhile, conventional ultrasonic cutting of amyloid fibers resulted in non-uniform fiber lengths that were difficult to control to the desired length. Furthermore, heat generated during the cutting process posed a risk of structural deformation of the amyloid fibrils, and there was a problem of fibers binding together after ultrasonic treatment.

[0040] Meanwhile, the inventors confirmed through experiments that when a pressure cutting process is repeated using a filter of a predetermined size and an extruder, and when the concentration of amyloid long fibers is controlled during the pressure cutting process, relatively long amyloid fibers can be obtained in the form of desired uniform short fibers, and that mass production is possible because the process is simple and inexpensive, thereby completing the present invention.

[0041] Hereinafter, a method for controlling the length of amyloid fibrils through a pressure cutting process according to the present invention will be described in more detail.

[0042]

[0043] Methods for controlling the length of amyloid fibrils

[0044] A method for controlling the length of amyloid fibrils according to one embodiment of the present invention may include: a) a step of synthesizing amyloid long fibers; and b) a step of extruding the synthesized amyloid long fibers and cutting them into amyloid short fibers of a predetermined length by passing them through a filter having a plurality of pores.

[0045]

[0046] First, synthesize amyloid intestinal fibers (step a).

[0047] Amyloid refers to the result of water-soluble proteins being converted into water-insoluble fibrous structures through selective and specific interactions between proteins under various biological and chemical conditions. Generally, it exhibits a lattice-shaped beta-sheet structure, in which hydrogen bonds between the protein backbones and the constituent proteins of the fiber are arranged perpendicular to each other. This allows for the formation of protein nanofibers with a width of approximately 2 to 20 nm that exhibit high strength comparable to that of a spider silk.

[0048] Meanwhile, in the present invention, the amyloid fiber is a long fiber, and in the present invention, the term long fiber may mean a fiber with a length of 800 nm or more.

[0049] The amyloid intestinal fiber according to one embodiment of the present invention may be formed by forming a fiber aggregate of one protein selected from egg white lysozyme (HEWL), Abeta42, Tau(AcPHF6), alpha-synuclein, insulin, and combinations thereof.

[0050] As a specific example, a method for synthesizing amyloid intestinal fibers formed by egg white lysozyme protein into fiber aggregates can be prepared by first separating and purifying egg white lysozyme, heating a solution in which the purified proteins are dissolved to a temperature range of 60 to 90°C under conditions of a pH range of 2 to 7, and then maintaining it at a predetermined temperature to produce a fiber shape.

[0051]

[0052] Next, the synthesized amyloid long fibers are extruded and cut into amyloid short fibers of a predetermined length by passing them through a filter having a plurality of pores (step b).

[0053] Step b above is performed by mechanically cutting the amyloid long fibers obtained in Step a in a top-down manner. Specifically, Step b can be performed by extruding the amyloid long fibers and passing them through a filter equipped with multiple pores. Meanwhile, Step b can be performed by controlling the length of the short fibers by controlling one or more of the number of extrusions, the filter pore size, and the amyloid long fiber concentration, as described below.

[0054] The filter used in the present invention has one material selected from polycarbonate, anodic aluminum oxide, polyethersulfone, and combinations thereof, and may include a plurality of pores having a diameter corresponding to the desired short fiber length.

[0055] Specifically, polycarbonate filters have very uniform pore sizes and low protein adsorption, making them suitable for controlling amyloid fiber length. On the other hand, while anodized aluminum filters allow for precise pore size control, their durability is relatively weak, which may lead to reduced durability during repetitive extrusion processes. Meanwhile, polyethersulfone filters have low protein adsorption and high chemical stability, but their pore size uniformity may be lower than that of polycarbonate filters.

[0056] In addition, polytetrafluoroethylene filters, which can be considered, have high chemical resistance, but their protein adsorption characteristics may vary depending on the specific study. Nylon filters have excellent mechanical strength, but their high protein adsorption may make them unsuitable for amyloid fiber research. Furthermore, polyvinylidene fluoride filters are suitable for protein purification, but precise control of pore size may be difficult.

[0057] Accordingly, the filter of the present invention may select and use a polycarbonate filter having high mechanical strength and excellent uniformity. The polycarbonate filter maintains its pore size even during a repetitive extrusion process and has a low degree of protein adsorption, making it most suitable for precisely cutting amyloid fibers.

[0058] Meanwhile, the pores of the filter may have a diameter in the range of 100 to 400 nm, specifically in the range of 100 to 200 nm. Specifically, when pressure is applied to a solution containing amyloid long fibers in the above step and the solution is passed through the filter, the amyloid long fibers are adsorbed onto the filter surface due to the pressure applied to the amyloid long fibers and the solution containing them, and subsequently, the long fibers are cut to a length corresponding to the pore size by the pressure. Meanwhile, in the present invention, the term "short fiber" may refer to a fiber having a length within the range of 100 to 400 nm.

[0059] Meanwhile, the extrusion process of step b above can be performed using, for example, a liposome extruder (Avanti, etc.), and the conditions for extrusion can be performed at a pressure in the range of 100 to 500 psi and an extrusion speed in the range of 2 to 5 mL / min.

[0060] Meanwhile, as the number of repetitions of the above step b increases, that is, as it is repeated, the length of the cut short fiber can converge to the pore diameter; therefore, it can be repeated, for example, 1 to 120 times (cycles), more specifically 80 to 100 times, and even more specifically 100 times. If the number of repetitions of the above step b is less than 80, the amyloid long fibers may not be sufficiently cut, making it difficult to obtain short fibers of the desired length, and there is a possibility that a large variation in length may occur as some long fibers pass through the filter. On the other hand, if the number of repetitions exceeds 120, the length of the short fibers may become excessively short due to excessive cutting, and the reproducibility of the experiment may decrease as fiber aggregation increases along with structural deformation of the original fibers. On the other hand, when the number of repetitions is 100, short fibers of the most uniform length can be secured, and accordingly, a stable and highly reproducible amyloid fiber cutting process can be effectively implemented.

[0061] In addition, during the extrusion of step b above, the concentration of amyloid fibers in the solution may be within the range of 0.1 to 2 wt% based on the total weight of the solution containing amyloid fibers. Meanwhile, if the concentration of amyloid fibers is below the above range, the applied pressure is reduced, making it difficult to obtain short fibers of uniform length. Specifically, if the concentration is less than 0.1 wt%, the viscosity of the solution decreases, resulting in insufficient pressure applied during extrusion; consequently, it may be difficult to obtain short fibers of uniform length, and there is a high probability that some long fibers will pass through the filter without being completely cut. On the other hand, if the concentration exceeds 2 wt%, aggregation between amyloid fibers increases, potentially clogging the filter pores. This may lead to a decrease in extrusion efficiency and a larger variation in the length of the cut fibers. Meanwhile, at a concentration of 1.0 wt%, stable and uniform short fibers can be secured, thereby optimizing the reproducibility of the experiment and the cutting efficiency.

[0062] In addition, step b above can be performed under constant temperature conditions while the amyloid long fibers are extruded. Specifically, the temperature of the entire extrusion system or the solution containing the fibers can be maintained in the range of 20 to 25°C so that the amyloid long fibers maintain a structurally stable state and efficient cutting is achieved during the extrusion process. This temperature range is effective in preventing structural deformation or unintentional re-aggregation of the amyloid short fibers and can contribute to securing short fibers of a uniform shape, particularly for precise analysis such as atomic force microscopy (AFM). Such temperature control can play an important role in improving the reproducibility and quality of the result by minimizing deviations in the length and diameter of the cut amyloid fibers.

[0063] Meanwhile, the amyloid short fibers prepared according to the pressure cutting method process including steps a and b above may have a loss rate of 20% or less based on the original fiber.

[0064]

[0065] Length-modulated amyloid short fibers

[0066] According to another embodiment of the present invention, the length-controlled amyloid fibers prepared according to the method may have a standard deviation of length in the range of 18 to 23% based on atomic force microscopy (AFM) analysis of 300 or more individual amyloid fibers.

[0067] Specifically, if the standard deviation of the amyloid fiber length exceeds 23%, the uniformity of the single fiber length decreases, making it difficult to obtain consistent results in experiments and applications, and there may be limitations in using it for precise analysis such as in research on neurodegenerative diseases or biosensors.

[0068] Meanwhile, the length-controlled amyloid fibers prepared according to the present invention can be effectively utilized in one or more technical fields selected from neurodegenerative disease research, biosensors, filtration systems, nanoelectronic devices, nanoparticles for drug screening, and bioadhesives.

[0069]

[0070] Examples

[0071] The present invention is capable of various modifications and may take various forms, and specific embodiments are illustrated and described in detail below. However, this is not intended to limit the invention to the specific disclosed forms, and it should be understood that the invention includes all modifications, equivalents, and substitutions that fall within the spirit and scope of the invention.

[0072]

[0073] Preparation Example: Synthesis of Amyloid Nanofibers (ANF)

[0074] The amyloid nanofibers (ANF) used in the embodiments of the present invention were synthesized based on Hen Egg White Lysozyme (HEWL) and prepared through the following process.

[0075] First, egg white lysozyme protein was dissolved in distilled water and adjusted to a pH range of 2.0 to 7.0. Subsequently, the solution was heated at a temperature of 60 to 90°C and maintained for a certain period of time to induce protein aggregation, and reaction conditions were adjusted to optimize the formation of amyloid fibers. The formed amyloid long fibers were extracted only from the supernatant and purified into pure amyloid nanofibers (ANF). The amyloid nanofibers produced by this method were prepared to maintain their unique β-sheet structure while being adjustable to a desired length in the extrusion cutting process of the present invention.

[0076]

[0077] Example 1-1

[0078] Amyloid nanofibers were prepared according to the above preparation example, with the concentration of amyloid long fibers in the solution set to 1 wt%, and a liposome extruder from Avanti was prepared, and a polycarbonate filter was used as the membrane. At this time, the pore size of the polycarbonate filter was prepared to be 200 nm. Meanwhile, the pressure cutting process using the extruder and filter was performed at a pressure in the range of 100 to 500 psi and an extrusion speed in the range of 2 to 5 mL / min, and the number of pressure cutting process cycles was set to 100 (see Fig. 3).

[0079]

[0080] Examples 1-2, 1-3

[0081] The procedure was carried out in the same manner as in Example 1-1, except that the polycarbonate filter was prepared with a pore size of 100 nm (Example 1-2) and 400 nm (Example 1-3).

[0082]

[0083] Comparative Examples 1-1, 1-2

[0084] The procedure was carried out in the same manner as in Example 1-1, but the number of pressure cutting process cycles was set to 50 (Comparative Example 1-1) and 0 (Comparative Example 1-2).

[0085]

[0086] Comparative Example 2

[0087] The procedure was carried out in the same manner as Example 1-1, except that the concentration of amyloid intestinal fibers in the solution was 0.1 wt%.

[0088]

[0089] [Experiment 1: Atomic Force Microscopy (AFM) Analysis and Statistical Analysis]

[0090] Figures 4 and 5 show the results of atomic force microscopy analysis and statistical analysis of the diameter and length of the fabricated amyloid short fibers.

[0091] Referring to the experimental results in Fig. 4, it was confirmed that in Example 1-1, Comparative Example 1-1, and Comparative Example 1-2, as the number of cycles increased, the length of the manufactured amyloid short fibers became closer to the pore size.

[0092] Referring to the experimental results in Fig. 5, it was confirmed that in both Examples 1-1 and 1-2, the amyloid short fiber lengths were formed to a size similar to the respective filter pore sizes.

[0093]

[0094] [Experiment 2: Atomic Force Microscopy (AFM) Analysis and BCA Assay]

[0095] Referring to the experimental results in Figure 6, in order to verify the efficiency of producing amyloid short fibers during the pressure cutting extrusion process, the carbonate filter was analyzed via AMF after the production of amyloid short fibers. As a result, it was confirmed that the amyloid short fibers were adsorbed onto the carbonate filter used previously, and it was confirmed that the remaining fibers, which were cut from the amyloid fibers, were spread radially around the filter pores.

[0096] Meanwhile, in order to confirm the loss rate according to the pressure cutting method of the present invention, untreated amyloid fibers were prepared according to Examples 1-1, 1-2 and the Preparation Example, respectively, and the total protein amount was analyzed by comparing it with the Control (Preparation Example) through a BCA assay. As a result, it was confirmed that the loss rate was 17.06% in the case of Example 1-1 and 14.19% in the case of Example 1-2.

[0097]

[0098] [Experiment 3: Experiment on Binding and Aggregation Between Amyloid Fibers]

[0099] Figure 7 shows the experimental results of confirming the binding and aggregation between amyloid fibers by reacting each of the untreated amyloid fibers according to Examples 1-1, 1-2 and the Preparation Example at a temperature of 24°C for 2 weeks. According to the experiment, when the amyloid fibers according to the Preparation Example (control), Example 1-1, and Example 1-2 were reacted at 24°C for 2 weeks, the fibril diameter was approximately 2-2.2 nm in both the first and second weeks, which was almost similar. However, it was confirmed that the length of the ANF according to Example 1-2 grew by about 25 nm per week, with 124.4 nm in the first week and 152.4 nm in the second week. According to Example 1-1, the length of the ANF was 217.3 nm at week 1 and 233.6 nm at week 2, confirming that the length grew by approximately 16-17 nm per week. Through these results, it was confirmed that the fabricated ANF can grow again.

[0100]

[0101] [Experiment 4: Measurement of Effects According to Concentration]

[0102] Figure 8 shows the effect according to the concentration of amyloid long fibers in the solution. In the case of Example 1-1, it was confirmed that the cleavage of amyloid long fibers was effectively performed, but in the case of Comparative Example 2, it was confirmed that the cleavage was not performed uniformly due to the influence of the lowered pressure.

[0103]

[0104] The foregoing description is merely an illustrative explanation of the technical concept of the present invention, and those skilled in the art to which the present invention pertains will be able to make various modifications and variations within the scope of the essential characteristics of the present invention. Accordingly, the embodiments disclosed in the present invention are intended to explain, not limit, the technical concept of the present invention, and the scope of the technical concept of the present invention is not limited by these embodiments. The scope of protection of the present invention shall be interpreted by the claims below, and all technical concepts within an equivalent scope shall be interpreted as being included within the scope of rights of the present invention.

Claims

1. a) a step of synthesizing amyloid intestinal fibers; and b) a step of cutting the synthesized amyloid long fiber into amyloid short fibers of a predetermined length by extruding the synthesized amyloid long fiber and passing it through a filter having a plurality of pores; a method for controlling the length of amyloid fibrils, comprising:

2. In Paragraph 1, A method for controlling the length of amyloid fibrils, wherein the amyloid intestinal fibers are formed by forming fiber aggregates of one protein selected from egg white lysozyme (HEWL), Abeta42, Tau(AcPHF6), alpha-synuclein, insulin, and combinations thereof.

3. In Paragraph 1, The above step a is a method for controlling the length of amyloid fibrils, wherein the solution in which purified protein is dissolved is heated to a temperature range of 60 to 90°C under conditions of a pH range of 2 to 7, and then maintained at a predetermined temperature to form a fiber shape.

4. In Paragraph 1, A method for controlling the length of an amyloid fibrillary, wherein, in step b above, the length of a short fiber is controlled by controlling one or more of the number of extrusions, filter pore size, and amyloid long fiber concentration.

5. In Paragraph 1, A method for controlling the length of amyloid fibrils, wherein in step b above, the filter has a material selected from polycarbonate, anodic aluminum oxide, polyethersulfone, and combinations thereof.

6. In Paragraph 1, A method for controlling the length of amyloid fibrils, wherein in step b above, the pores of the filter have a diameter in the range of 100 to 400 nm.

7. In Paragraph 1, A method for controlling the length of amyloid fibrils, wherein the extrusion of step b above is performed at a pressure in the range of 100 to 500 psi and an extrusion speed in the range of 2 to 5 mL / min.

8. In Paragraph 1, A method for controlling the length of amyloid fibrils, wherein the extrusion of step b above is repeated 1 to 120 times.

9. In Paragraph 1, A method for controlling the length of amyloid fibrils, wherein the concentration of amyloid fibers extruded in step b above is within the range of 0.1 to 2 wt% based on the total weight of the solution containing amyloid fibers.

10. In Paragraph 1, A method for controlling the length of amyloid fibrils, wherein the extrusion of amyloid long fibers in step b above is performed under conditions maintained in a temperature range of 20 to 25 ℃.

11. In Paragraph 1, The amyloid short fiber prepared according to the pressure cutting method process including steps a and b above is an amyloid short fiber length control method having a short fiber reference loss rate of 20% or less.

12. Length-controlled amyloid fibers prepared according to the method of claim 1, The amyloid fibers are amyloid fibers in which the standard deviation of the length is within the range of 18 to 23% based on atomic force microscopy (AFM) analysis of 300 or more individual amyloid fibers.

13. In Paragraph 12, The amyloid fibers are used in one or more technical fields selected from neurodegenerative disease research, biosensors, filtration systems, nanoelectronic devices, nanoparticles for drug screening, and bioadhesives.

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