Phosphorylation-Dependent Protein Motif for Kinase Detection

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Solution Overview

Problem

Current methods for identifying and developing inhibitors for serine and threonine kinases, which play crucial roles in cellular and extracellular functions, are limited, as most kinase inhibitors target tyrosine kinases, and there is a need for new tools to interrogate and respond to the activities of protein kinases and phosphatases.

Innovation Solution

Designing a phosphorylation-dependent protein motif that undergoes structural changes based on its phosphorylation state by replacing structurally significant glutamic acid or aspartic acid residues with serine or threonine residues, allowing for the detection of protein kinase and phosphatase activities and the identification of inhibitors through phosphorylation or dephosphorylation events.

Engineering Contradictions & Design Principles

VSEngineering Contradiction Analysis

1Adaptability or versatility

If most kinase inhibitors target tyrosine kinases, then existing pharmaceutical development is well-established, but there is a lack of inhibitors for serine and threonine kinases

Engineering Contradiction:

Improvekinase inhibitor specificityVSAvoidinhibitor development reliability

Core Design Contradiction:

Adaptability or versatilityVSReliability

Solution Approach 1:

The patent changes the target parameter from tyrosine kinase to serine/threonine kinase inhibition. By modifying the kinase inhibitor to specifically target serine and threonine residues instead of tyrosine, the invention expands the versatility of kinase inhibition to cover previously underrepresented kinases, addressing the technical contradiction between established tyrosine kinase inhibitors and the need for serine/threonine kinase inhibitors

Inventive Principle:

Principle #35Parameter changes

2Adaptability or versatility

If new tools are needed to interrogate protein kinase and phosphatase activities, then research capability is improved, but current detection methods are insufficient

Engineering Contradiction:

Improvedetection tool versatilityVSAvoidkinase activity detection difficulty

Core Design Contradiction:

Adaptability or versatilityVSDifficulty of detecting and measuring

Solution Approach 1:

The patent introduces a phosphopeptide as an intermediary molecule that mediates between the kinase/phosphatase enzyme and the detection system. The phosphopeptide contains a fluorescent probe that becomes fluorescent upon phosphorylation, serving as an intermediary signal that translates invisible enzymatic activity into a detectable optical signal, thereby resolving the contradiction between needing versatile detection tools and the difficulty of measuring kinase activity

Inventive Principle:

Principle #24Intermediary (Mediator)

3Adaptability or versatility

If phosphorylation-dependent structural changes are designed, then protein function regulation is achieved, but structural design complexity increases

Engineering Contradiction:

Improveprotein function regulationVSAvoidprotein motif design complexity

Core Design Contradiction:

Adaptability or versatilityVSDevice complexity

Solution Approach 1:

The patent applies local quality by introducing a fluorescent probe at a specific local position within the peptide sequence rather than throughout the entire protein. The probe is positioned at a conserved site that becomes fluorescent upon phosphorylation, allowing the rest of the peptide to maintain its native structure and function while providing a localized signal. This resolves the contradiction between achieving phosphorylation-dependent regulation and avoiding excessive design complexity

Inventive Principle:

Principle #3Local quality

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 the effective identification of protein kinase and phosphatase activities and the screening of inhibitors, providing a novel approach to target serine and threonine kinases, which are essential for various cellular processes and disease treatments.

Implementation Method 1

replacing a structurally significant glutamic acid or aspartic acid residue in a polypeptide with a serine or threonine residue... activating the domain comprising the replacement serine or threonine residue by phosphorylating the serine or threonine residue

Methodology Applied

Scientific EffectPhosphorylation: Chemical Bonding

Implementation Method 2

identifying protein phosphatase activity by detecting dephosphorylation of the replaced phosphoserine or phosphothreonine residue

Methodology Applied

Scientific EffectDephosphorylation: Chemical Bonding

Data Source

PatentUS8148506B2Protein kinase-inducible domains

Publication Date: 2012.04.03 UNIVERSITY OF DELAWARE

AI summary

Applicants have used protein design to develop novel functional protein architectures, termed protein kinase-inducible domains, whose structures are dependent on phosphorylation by specific protein kinases or are dependent on dephosphorylation by specific protein phosphatases. Applicants have designed kinase-inducible domains based on a modular architecture, which allows kinase-inducible domains to be responsive to any specific serine-threonine kinases. Kinase-inducible domains can consist of canonical amino acids, allowing their use as expressible tags of protein kinase activity.