Protein Sequencing via DNA Barcode Transfer
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Solution Overview
Problem
Current protein and peptide sequencing technologies face limitations in throughput, accuracy, and sensitivity, particularly in handling large datasets and low-abundance proteins, with existing methods like Edman degradation and mass spectrometry being costly, requiring sophisticated users, and having limited dynamic range and sequencing length.
Innovation Solution
A method involving immobilizing polypeptides on solid surfaces with sample ID DNA tags, binding N-terminal amino acids with modification molecules and coding DNA tags, cyclically transferring and amplifying DNA barcodes, and decoding using hybridization assays or sequencing to achieve high parallelization and multiplexing.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If Edman degradation is used for peptide sequencing, then the sequencing process can be performed with established methodology, but the sequencing length is limited to under 30 residues in practice due to incomplete cyclic derivation
Solution Approach 1:
The patent introduces DNA barcodes as intermediary molecules that bind to N-terminal amino acids. Instead of directly sequencing the peptide through multiple Edman cycles, the method transfers amino acid information to stable DNA barcodes, which can then be sequenced using robust DNA sequencing technologies. This intermediary approach bypasses the limitation of incomplete cyclic derivation in traditional Edman degradation and enables sequencing of much longer peptides.
2Productivity
If mass spectrometry is used for protein sequencing, then sequencing can be performed with high speed and reduced cost, but the dynamic range is limited and low-abundance proteins cannot be effectively analyzed
Solution Approach 1:
The patent segments the protein sequencing problem into two independent parts: (1) enrichment and identification of target peptides through immunocapture with high specificity, and (2) parallel sequencing of many individual peptide molecules using DNA barcode transfer and sequencing. This segmentation allows each step to be optimized independently, enabling both high sensitivity for low-abundance proteins and high throughput for large-scale analysis.
3Productivity
If high throughput sequencing is implemented, then the analysis of large datasets and multiple samples can be performed, but the device complexity and requirement for sophisticated operation increase
Solution Approach 1:
The patent creates physical copies of peptide information in the form of DNA barcodes that can be amplified and sequenced. By transferring the sequential information from a single peptide molecule into a DNA barcode sequence, the method enables replication of the information across many copies, which can then be processed using standardized, relatively simple DNA sequencing instruments rather than requiring complex custom-built protein sequencing devices.
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
This approach enables efficient, accurate, and sensitive protein and peptide sequencing, overcoming limitations of existing methods by allowing for longer sequencing lengths and higher throughput, including the analysis of low-abundance proteins, within a single instrument.
Implementation Method 1
binding the N-terminal amino acid (NTAA) of the polypeptide with a modification molecule; contacting the polypeptide with a NTAA binding molecule
Implementation Method 2
amplifying the coding DNA strands into a cluster in situ on solid surfaces
Implementation Method 3
decoding DNA strands on the solid surfaces using hybridization assays or sequencing
Data Source
AI summary
This disclosure is directed to a method for sequencing protein and peptide. The method specifies the steps of providing a polypeptide that is immobilized on solid surfaces and binding a N-terminal amino acid (NTAA) of the polypeptide with a modification molecule. Further, the method requires contacting the polypeptide with a NTAA binding molecule with a DNA coding tag, transferring an information from the DNA coding tag of the NTAA binding molecule to a universal primer and then forming an extended DNA tag on the solid surfaces, and cleaving the N-terminal amino acid on the polypeptide. Then, step (b) through step (e) are cyclically repeated. Finally, extended DNA strains are decoding on the solid surfaces.