Single-molecule peptide sequencing via barcode transfer reagent
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Solution Overview
Problem
Current single-molecule protein sequencing methods face inefficiencies due to the small size and internal sequestration of amino acids within proteins, making it difficult for bulky identification tools to access and label them effectively.
Innovation Solution
The method involves coupling peptides with barcodes and using a Barcode Transfer Reagent (BTR) to generate barcoded-amino acid complexes, which are then cleaved and read out to sequence terminal amino acids, overcoming the limitations of bulky tools and internally sequestered amino acids by separating and identifying amino acids ex-situ.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If bulky identification tools are used to label amino acids within proteins, then amino acid identification can be performed, but the tools cannot effectively access and label internally sequestered amino acids
Solution Approach 1:
The patent extracts amino acids from their native protein environment by performing proteolytic digestion to release individual amino acids or small peptides. These extracted amino acids are then barcoded ex-situ using standardized reagents, eliminating the accessibility problem of bulky tools while maintaining identification accuracy through controlled extraction and sequencing processes
Solution Approach 2:
The patent segments the protein into individual amino acids or small peptide fragments through proteolytic digestion. This segmentation transforms the inaccessibility problem of internally sequestered amino acids into manageable units that can be individually barcoded and sequenced, with the original sequence information recovered through computational assembly of the barcoded fragments
2Productivity
If amino acids are identified within their native protein environment, then structural context is preserved, but identification efficiency is reduced due to accessibility limitations
Solution Approach 1:
The patent introduces barcodes as intermediary molecules that bridge the amino acid sequence information and the sequencing platform. Each amino acid or peptide fragment receives a unique barcode that serves as a mediator, carrying the identity information through the ex-situ barcode transfer and amplification processes to the sequencing step, thereby enabling high-throughput processing while maintaining sequence assembly accuracy through the faithful mediation of barcode information
Solution Approach 2:
The patent creates copies of amino acid identity information in the form of barcodes that can be amplified and sequenced independently of the original protein structure. The barcode copying process allows multiple copies of the same amino acid identity to be generated and detected, increasing throughput while the original sequence information is preserved through the copying fidelity of the barcode transfer reaction
Data Source
AI summary
The present disclosure provides systems and methods directed towards single molecule protein sequencing, through use of a barcode transfer reagent. Systems and methods described herein allow for massively parallel single-molecule protein sequencing through molecular barcoding and ex-situ analysis.