VEGF-Binding Aptamer Affinity Enhancement via In Silico Maturation
Find Innovative SolutionsGenerate Solutions
Solution Overview
Problem
Current methods for measuring VEGF, such as immunoassays using antibodies, are laborious and expensive, and existing aptamers have limited sensitivity due to low affinity and secondary structure issues during PCR amplification in the SELEX process.
Innovation Solution
The development of a novel VEGF-binding aptamer with enhanced affinity through in silico maturation, where the base sequence of VEap121 is mutated to improve binding capacity, and the creation of a bivalent aptamer by linking two 3R02 molecules via a linker to increase binding efficiency.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If conventional SELEX method is used to prepare VEGF-binding aptamer, then aptamer can be obtained through chemical synthesis, but the affinity to VEGF is insufficient (dissociation constant higher than 3 nM)
Solution Approach 1:
The patent applies parameter changes by modifying the base sequence of the aptamer through in silico maturation. Specifically, the base sequence was mutated to optimize binding capacity, transforming the original SELEX-obtained sequence into an improved sequence with dissociation constant less than 3 nM. This sequence optimization directly enhances binding affinity and detection sensitivity.
Solution Approach 2:
The patent creates a bivalent aptamer by linking two 3R02 aptamer molecules via a linker sequence. This composite structure combines multiple binding units into a single molecule, significantly enhancing the overall binding affinity to VEGF and achieving a dissociation constant of less than 3 nM, thereby resolving the insufficient affinity problem of conventional aptamers.
2Measurement precision
If immunoassay using antibodies is used to measure VEGF, then specific detection can be achieved, but the process becomes laborious and expensive
Solution Approach 1:
The patent replaces expensive and laborious antibody-based immunoassays with aptamers that can be chemically synthesized using automated nucleic acid synthesizers. The aptamer sequence is designed to be stable and functional, eliminating the need for complex antibody production processes while maintaining detection specificity, thereby significantly reducing production complexity and cost.
Solution Approach 2:
The patent substitutes the biological system of antibody production and purification with a chemical synthesis system. The aptamer's nucleic acid structure allows for automated chemical synthesis, replacing the mechanical and biological processes required for antibody manufacturing, thus simplifying production while maintaining detection capability.
3Measurement precision
If aptamer with high affinity to VEGF is developed, then detection sensitivity is improved, but the secondary structure may interfere with PCR amplification during SELEX process
Solution Approach 1:
The patent applies preliminary action by performing in silico maturation and sequence optimization before final aptamer synthesis and application. The base sequence was pre-optimized through computational methods to ensure high binding affinity while maintaining amplification compatibility, preventing secondary structure interference issues before they arise in the actual SELEX and PCR processes.
Applied Scientific Principles
This section explains which scientific principles are used to turn an abstract innovation direction into a practical engineering solution.
Function Achieved in This Case
The novel aptamer achieves a significantly higher affinity to VEGF, with a dissociation constant of less than 3 nM, enabling highly sensitive measurement of VEGF in test samples, improving detection and quantification sensitivity compared to previous aptamers.
Implementation Method 1
an aptamer that binds to vascular endothelial growth factor (VEGF) with a dissociation constant of less than 3 nM
Data Source
AI summary
Disclosed is providing a novel VEGF-binding aptamer whose affinity to VEGF is higher than those of known VEGF-binding aptamers. By an in silico maturation method starting from a known VEGF-binding aptamer, 4 kinds of aptamers whose affinities to VEGF are higher than that of the known VEGF-binding aptamer were prepared. By linking two molecules of an obtained aptamer to each other via a linker, an aptamer having an even higher affinity to VEGF was obtained. The polynucleotide of the present invention contains the base sequence of any one of SEQ ID NOs:1 to 4, and binds to vascular endothelial growth factor.