Compound Protease Logic Circuits in Living Cells
Find Innovative SolutionsGenerate Solutions
Solution Overview
Problem
Current synthetic biology approaches face limitations in designing new functions in living cells, particularly in implementing protein-level circuits that can effectively regulate protein activity, localization, and stability without modifying the genome or entering the nucleus.
Innovation Solution
A compound protease is developed, comprising a protease domain and a cut site, where the protease domain forms an active form only when its parts associate together, and an association domain that stabilizes this active form, allowing for controlled activation and deactivation by specific enzymes, enabling the creation of synthetic protein circuits for logic gates and other complex cellular functions.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Adaptability or versatility
If gene regulation circuits are used to implement cellular functions, then the design of new functions in living cells is limited, but the complexity of protein-level circuits is reduced
Solution Approach 1:
The patent replaces gene-level regulation with protein-level circuits, substituting transcriptional control mechanisms with direct protein-protein interactions and proteolytic events. This enables faster response times and more sophisticated logic operations while maintaining manageable complexity through modular protease design
Solution Approach 2:
The protease molecules are divided into separate domains (protease domain, association domain, cut site) that can independently function and recombine. This segmentation allows for modular assembly of complex circuits from standardized parts, increasing design versatility without proportionally increasing overall system complexity
2Adaptability or versatility
If protease molecules are designed with multiple domains and cut sites, then the ability to perform logic operations is enhanced, but the structural complexity of the protease increases
Solution Approach 1:
The protease is designed with multiple functional domains that serve different purposes: the protease domain performs catalysis, the association domain mediates protein-protein interactions, and the cut site enables regulated cleavage. This multi-functionality allows a single protease molecule to participate in complex logic operations while maintaining a standardized structural framework that can be reused across different circuit designs
3Reliability
If the protease domain parts do not self-associate, then the control over protease activation is improved, but the stability of the inactive protease parts decreases
Solution Approach 1:
The association domain acts as an intermediary that bridges the separated protease domain parts. In the inactive state, the association domain maintains the protease parts in a stable, non-self-associating configuration. Upon receiving the appropriate signal, the association domain facilitates controlled association of the protease parts, enabling precise activation control while maintaining structural stability in both active and inactive states
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
This solution enables programmable protein circuits that can perform logic operations, regulate cellular behaviors, and control cellular processes like apoptosis, offering a versatile and scalable method for implementing various functions without genomic integration.
Implementation Method 1
a cut site, wherein the cut site comprises: a first part of the cut site, wherein the first part of the cut site is linked to the first part of the protease domain; and a second part of the cut site, wherein the second part of the cut site is linked to the second part of the protease domain, wherein when the first and second parts of the cut site are associated together they form an active cut site for an enzyme, and wherein when the active cut site is cut by the enzyme, the first and second parts of the cut site dissociate from one another
Implementation Method 2
an association domain, the association domain comprising: a first part of the association domain that is conjugated to the second part of the cut site; a second part of the association domain that is linked to the second part of the protease domain, wherein the association domain is configured to stabilize the active protease domain
Data Source
AI summary
The present disclosure relates to systems, methods and compositions provided herein that include a compound protease. The compound protease can contain a protease domain, a cut site for another enzyme and an association domain. In some embodiments, the compound protease is part of a protein circuit.