Glycosylated YghJ Polypeptides Enrichment via BEMAP

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Solution Overview

Problem

Current methods for understanding and detecting bacterial glycoproteins, particularly in Enterotoxigenic Escherichia coli (ETEC), face challenges due to limited analytical tools, heterogeneous glycosylation, low abundance, and poor ionization of glycosylated peptides, hindering the discovery of novel therapeutic targets and their immunogenic potential.

Innovation Solution

Development of a mass spectrometry-based technique called BEMAP (Bacterial Enrichment through Michael Addition and Phosphorylation), which substitutes O-linked carbohydrate moieties with 2-Aminoethyl phosphonic acid (AEP), allowing for selective enrichment and improved fragmentation analysis of glycosylated peptides, enabling the identification of immunogenic glycosylated YghJ polypeptides.

Engineering Contradictions & Design Principles

VSEngineering Contradiction Analysis

1Measurement precision

If conventional analytical tools are used to detect bacterial glycoproteins, then the detection process is simple, but the detection precision and ability to identify glycosylated peptides is poor due to heterogeneous glycosylation, low abundance, and poor ionization

Engineering Contradiction:

Improvedetection precisionVSAvoidanalytical tool complexity

Core Design Contradiction:

Measurement precisionVSDevice complexity

Solution Approach 1:

The patent introduces an intermediary chemical reaction (Michael addition) that substitutes O-linked carbohydrate moieties with 2-Aminoethyl phosphonic acid (AEP). This intermediary step converts poorly ionizing glycosylated peptides into forms with improved ionization and detectability, resolving the contradiction between detection precision and analytical tool complexity

Inventive Principle:

Principle #24Intermediary (Mediator)

Solution Approach 2:

The patent changes the chemical parameters of the glycosylated peptides by substituting carbohydrate moieties with AEP groups. This parameter change improves ionization efficiency and detectability without requiring complex analytical instruments, thereby improving measurement precision while avoiding increased device complexity

Inventive Principle:

Principle #35Parameter changes

2Productivity

If existing enrichment methods are used, then the process is straightforward, but the enrichment efficiency is low due to limited specificity for bacterial glycosylated proteins

Engineering Contradiction:

Improveenrichment efficiencyVSAvoidmethod complexity

Core Design Contradiction:

ProductivityVSEase of manufacture

Solution Approach 1:

The patent extracts and removes the O-linked carbohydrate moieties from bacterial glycoproteins and replaces them with AEP groups. This extraction approach specifically targets glycosylated peptides while leaving non-glycosylated peptides unchanged, achieving high enrichment efficiency with a relatively simple chemical approach

Inventive Principle:

Principle #2Taking out (Extraction)

Solution Approach 2:

The patent converts the harmful effect of heterogeneous glycosylation (which obscures peptide identification) into a beneficial feature by using the glycan moieties themselves as targets for substitution. The glycosylation that previously hindered detection is now the very feature being exploited for selective enrichment and identification

Inventive Principle:

Principle #22Blessing in disguise (Convert harm into benefit)

3Loss of information

If glycosylated peptides are analyzed directly, then no additional processing is needed, but the immunogenicity assessment is inaccurate due to poor ionization and low abundance

Engineering Contradiction:

Improveinformation lossVSAvoidprocessing time

Core Design Contradiction:

Loss of informationVSLoss of time

Solution Approach 1:

The patent performs preliminary chemical modification (Michael addition substitution) on the glycosylated peptides before analysis. This preliminary action improves ionization and detectability, ensuring that immunogenicity information is not lost during subsequent mass spectrometry analysis, while the streamlined process minimizes additional time investment

Inventive Principle:

Principle #10Preliminary action

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

BEMAP effectively maps and enriches glycosylated peptides, revealing enhanced immunogenicity of YghJ proteins, thereby providing a novel approach for vaccine development and therapeutic targeting of ETEC infections.

Implementation Method 1

BEMAP (Bacterial Enrichment through Michael Addition and Phosphorylation), which substitutes O-linked carbohydrate moieties with 2-Aminoethyl phosphonic acid (AEP)

Methodology Applied

Scientific EffectMichael addition: Chemical Bonding

Implementation Method 2

BEMAP (Bacterial Enrichment through Michael Addition and Phosphorylation), which substitutes O-linked carbohydrate moieties with 2-Aminoethyl phosphonic acid (AEP)

Methodology Applied

Scientific EffectPhosphorylation: Chemical Bonding

Data Source

PatentUS11655274B2Glycosylated YghJ polypeptides from enterotoxigenic <i>Escherichia coli </i>(ETEC)

Publication Date: 2023.05.23 AARHUS UNIV

AI summary

The present invention relates to glycosylated YghJ polypeptides from or derived from enterotoxigenic Escherichia coli (ETEC) that are immunogenic. In particular, the present invention relates to compositions or vaccines comprising the polypeptides and their application in immunization, vaccination, treatment and diagnosis of ETEC.