Multivalent Chelator Scaffold for Stable Protein Binding

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Solution Overview

Problem

Current chelator compounds used for modifying and immobilizing proteins, such as those with histidine-tags, suffer from instability and low binding affinity, making it difficult to achieve stoichiometrically defined and reversible interactions, especially in solution or within living cells.

Innovation Solution

Development of multivalent chelator compounds with multiple metal-coordinative centers, such as bis-NTA, tris-NTA, and tetrakis-NTA, attached to a scaffold structure, which provide a stable and reversible binding to affinity-tags like histidine-tags, allowing for the attachment of various probes and functional units.

Engineering Contradictions & Design Principles

VSEngineering Contradiction Analysis

1Reliability

If monovalent chelators (e.g., NTA) are used for protein modification, then the binding is reversible and easy to manipulate, but the binding stability and affinity are insufficient

Engineering Contradiction:

Improvebinding stabilityVSAvoidchelator structure complexity

Core Design Contradiction:

ReliabilityVSDevice complexity

Solution Approach 1:

The patent combines multiple chelator units (nitrilotriacetic acid groups) into a single multivalent chelator molecule that can simultaneously coordinate with multiple metal ions. This merging of multiple binding sites into one molecule enables stable, stoichiometric binding to histidine-tags while maintaining reversibility through metal ion competition, resolving the contradiction between binding stability and structural simplicity.

Inventive Principle:

Principle #5Merging (Combining)

Solution Approach 2:

The invention creates composite chelator structures by attaching multiple NTA groups to a central scaffold (such as glucose or other core structures). This composite approach allows the chelator to present multiple metal-coordinating sites in a controlled geometry, achieving high binding affinity and stability while maintaining a defined molecular structure that facilitates reversible binding.

Inventive Principle:

Principle #40Composite materials

2Reliability

If high density of NTA-groups is used in affinity matrix, then binding stability is improved, but the binding cannot be stoichiometrically defined at molecular level

Engineering Contradiction:

Improvebinding stabilityVSAvoidstoichiometric definition

Core Design Contradiction:

ReliabilityVSManufacturing precision

Solution Approach 1:

The patent segments the binding function into discrete, defined molecular units. Each multivalent chelator molecule contains a specific number of NTA groups (e.g., 2-4) attached to a defined scaffold, creating a segmented structure with precise stoichiometry. This allows each chelator-protein complex to have a well-defined stoichiometric ratio, unlike the continuous density variations in high-density NTA matrices.

Inventive Principle:

Principle #1Segmentation

Solution Approach 2:

The invention changes the valency parameter of the chelator from monovalent (single NTA group) to multivalent (multiple NTA groups per molecule). This parameter change enables the chelator to achieve high binding stability through multiple simultaneous metal ion coordination events while maintaining a defined molecular structure that permits stoichiometric characterization of the protein-chelator complex.

Inventive Principle:

Principle #35Parameter changes

3Reliability

If extended histidine-tags are used, then binding stability to metal-chelator is improved, but stoichiometrically defined and stable complexes cannot be achieved

Engineering Contradiction:

Improvebinding stabilityVSAvoidcomplex stoichiometry

Core Design Contradiction:

ReliabilityVSManufacturing precision

Solution Approach 1:

Instead of extending the histidine-tag length to improve binding stability, the patent inverts the approach by keeping the histidine-tag at standard length (6-10 residues) and instead extending the chelator valency. The multivalent chelator with multiple NTA groups provides enhanced stability through multiple coordination sites while maintaining a defined 1:1 complex stoichiometry between the multivalent chelator and the protein, avoiding the stoichiometric complexity that would arise from multiple histidine-tags.

Inventive Principle:

Principle #13The other way round (Inversion)

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 multivalent chelators enable stable, stoichiometric, and reversible interactions with proteins, facilitating selective and position-specific modification, immobilization, and detection, with enhanced stability and versatility compared to traditional monovalent chelators.

Implementation Method 1

multivalent chelator compounds with multiple metal-coordinative centers, such as bis-NTA, tris-NTA, and tetrakis-NTA

Methodology Applied

Scientific EffectMetal coordination:

Implementation Method 2

CL is a chelator-group with at least a metal-coordinative centre

Methodology Applied

Scientific EffectChelation:

Data Source

PatentUS9606114B2Multivalent chelators containing a scaffold structure for modifying and organizing of target molecules

Publication Date: 2017.03.28 JOHANN WOLFGANG GOETHE UNIV FRANKFURT AM MAIN

AI summary

New compounds of the general formulaXm-G-CLn are described as well as methods for their production and use in the analysis, detection and purification of target molecules. These constitute multivalent chelator-compounds with an affinity-tag binding to metal-chelator-complexes which can selectively modify and/or immobilize target molecules by a multitude of probes or functional units.