CRISPR-Transposon Protein Complexes for Targeted DNA Integration

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Solution Overview

Problem

Existing CRISPR systems lack efficient nucleic acid integration and modification capabilities, particularly in non-canonical pathways where nuclease components are absent or repurposed for alternative functions.

Innovation Solution

Engineered polypeptides and nucleic acids, including transposon-associated proteins like TnsA, TnsB, TnsC, and TniQ, and Cas proteins such as Cas5, Cas6, Cas7, and Cas8, are developed to enhance nucleic acid integration and binding activities, leveraging CRISPR-associated transposon (CRISPR-Tn) systems.

Engineering Contradictions & Design Principles

VSEngineering Contradiction Analysis

1Reliability

If conventional CRISPR systems are used, then basic immune function is provided, but nucleic acid integration capability is insufficient

Engineering Contradiction:

Improvenucleic acid integration capabilityVSAvoidintegration efficiency

Core Design Contradiction:

ReliabilityVSProductivity

Solution Approach 1:

The patent combines CRISPR systems with transposon components (Tns proteins) to create a hybrid system that merges the targeting precision of CRISPR with the integration efficiency of transposons. The engineered polypeptides include both Cas proteins and Tns proteins that work together to achieve both reliable targeting and efficient nucleic acid integration.

Inventive Principle:

Principle #5Merging (Combining)

Solution Approach 2:

The invention creates composite protein complexes by fusing or co-expressing CRISPR-associated proteins with transposon proteins. These composite polypeptides integrate the functional domains of both systems, resulting in a unified molecular machine that performs both guide RNA-directed targeting and transposon-mediated integration.

Inventive Principle:

Principle #40Composite materials

2Adaptability or versatility

If nuclease components are removed for non-canonical pathways, then alternative functions are achieved, but integration activity is reduced

Engineering Contradiction:

Improvealternative function capabilityVSAvoidnucleic acid integration activity

Core Design Contradiction:

Adaptability or versatilityVSReliability

Solution Approach 1:

The engineered polypeptides are designed to perform multiple functions within a single protein complex. The system can switch between different modes of action depending on the presence of guide RNA and other regulatory elements, enabling both transcriptional repression and nucleic acid integration without requiring separate systems.

Inventive Principle:

Principle #6Universality (Multi-functionality)

Solution Approach 2:

The patent introduces localized modifications to specific domains of the protein complexes to enhance integration activity in particular regions. The Tns proteins are engineered with specific amino acid substitutions that create localized functional enhancements at the integration site while maintaining overall system versatility.

Inventive Principle:

Principle #3Local quality

Data Source

PatentUS20250376701A1Crispr-transposon systems and components

Publication Date: 2025.12.11 THE BROAD INST INC

AI summary

The present disclosure provides Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)-associated transposon (CRISPR-Tn or CAST) systems, components thereof, and methods for nucleic acid modification using the systems or components. More particularly, the disclosure provides modified Cas proteins and transposon-associated proteins for nucleic acid modification.