Lectin Pre-Enrichment for Multiplexed PTM Protein Analysis
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Solution Overview
Problem
Existing methods are inadequate for quickly and inexpensively measuring multiple post-translational modifications of proteins in a protein-specific manner.
Innovation Solution
A method involving pre-enrichment of post-translationally modified proteins using lectins that specifically bind to saccharides in PTMs, followed by separation and detection using labeled binding molecules that target specific epitopes of the proteins.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Adaptability or versatility
If conventional methods are used to measure multiple post-translational modifications, then measurement capability is limited, but cost and time consumption increase
Solution Approach 1:
The assay is divided into distinct functional modules: lectin-based pre-enrichment module for capturing glycosylated proteins, binding molecule detection module for specific epitope recognition, and signal detection module for quantification. This segmentation allows each module to be optimized independently while enabling rapid multiplexed measurement of multiple PTMs
Solution Approach 2:
The lectin pre-enrichment step is performed beforehand to selectively capture and concentrate glycosylated proteins from complex samples. This preliminary action simplifies subsequent detection steps and enables rapid analysis of multiple PTMs by pre-sorting target proteins based on their glycosylation patterns
2Adaptability or versatility
If conventional methods are used to measure multiple post-translational modifications, then measurement capability is limited, but cost increases
Solution Approach 1:
The assay uses universal lectin reagents that can recognize multiple types of glycosylated proteins across different samples and disease states. The same lectin panel can be used to detect various PTMs on different target proteins, reducing the need for multiple specialized assays and lowering overall measurement costs
Solution Approach 2:
The method employs binding molecules with labels that can be detected through standardized readout systems. By using copyable detection signals (such as fluorescent or colorimetric labels), the assay enables high-throughput measurement of multiple PTMs using common instrumentation, reducing per-sample costs
3Measurement precision
If specific detection of multiple PTMs is performed, then measurement precision is improved, but device complexity increases
Solution Approach 1:
Lectins serve as intermediary reagents that specifically recognize and bind to glycosylated proteins through their carbohydrate moieties. This intermediary step provides high specificity for enriching glycosylated proteins while keeping the overall assay design simple and manageable
Solution Approach 2:
The assay employs a nested structure where binding molecules are embedded within lectin-protein complexes. The lectin captures the glycosylated protein first, then the binding molecule specifically binds to an epitope on the captured protein, creating a nested recognition system that enhances detection precision without requiring simultaneous complex interactions
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
Enables efficient quantification and identification of multiple PTMs in proteins, providing a cost-effective and rapid analysis of protein modifications.
Implementation Method 1
contacting the sample or a subsample thereof with a first lectin that specifically binds a first saccharide present in a post-translational modification (PTM) on one or more target proteins
Implementation Method 2
contacting the first pre-enriched subsample with a plurality of binding molecules comprising a first binding molecule that specifically binds a first epitope of a first target protein and a second binding molecule that specifically binds a second epitope of the first target protein
Data Source
AI summary
Provided herein are methods of analyzing post-translationally modified proteins. The methods may comprise pre-enriching proteins using lectins and/or multiplexed detection of multiple proteins and/or multiple post-translational modifications. Provided herein are also methods for determining the likelihood that a subject has a disease or condition, such as cancer.