De Novo Binding Polypeptides Without Disulfide Pairing Bottlenecks
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Solution Overview
Problem
Existing antibody-based reagents are costly and time-consuming, and alternative affinity reagents with improved specificity and affinity are needed to address these issues, particularly for therapeutic and diagnostic applications.
Innovation Solution
Development of de novo binding domain (DBD) polypeptides with specific amino acid sequences that provide high target binding affinity and specificity, including methods for producing and screening these polypeptides, as well as their use in therapeutic and diagnostic applications.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If disulfide bonds are introduced to constrain peptide structure, then binding affinity and specificity are improved, but improper cysteine pairing occurs leading to lower expression and product quality
Solution Approach 1:
The invention extracts and removes the problematic disulfide bond constraint from the peptide structure. By designing constrained peptides without disulfide bonds, the patent eliminates the source of improper cysteine pairing while maintaining structural constraint through alternative means (cyclic structure formed by backbone connectivity), thereby resolving the contradiction between binding affinity and expression yield
Solution Approach 2:
The invention changes the structural parameters of the peptide by defining specific constraints on the backbone chain (cyclic structure) without introducing disulfide bonds. This parameter change allows the peptide to maintain a fixed three-dimensional structure for high affinity binding while avoiding the manufacturing problems associated with cysteine pairing
2Reliability
If disulfide bonds are introduced to constrain peptide structure, then binding specificity is improved, but product quality deteriorates due to improper pairing
Solution Approach 1:
The invention extracts and removes the disulfide bond from the molecular structure, eliminating the source of improper pairing that compromises product quality. The peptide maintains its binding specificity through the constrained cyclic structure formed by the backbone chain, achieving high specificity without the quality issues of disulfide-based constraints
3Reliability
If traditional antibody-based reagents are used, then high binding affinity is achieved, but production costs and time increase
Solution Approach 1:
The invention employs simple, small peptide molecules as alternative affinity reagents to traditional antibodies. These peptides are cheaper to produce and can be synthesized more efficiently, providing a cost-effective substitute that maintains binding functionality while improving production efficiency
Solution Approach 2:
The invention changes the scale and complexity parameters by using small constrained peptides instead of large antibody molecules. This parameter change enables more efficient production through chemical synthesis or expression in simpler systems, reducing both cost and time while maintaining the essential binding function
Data Source
AI summary
Provided herein are de novo binding domain containing polypeptides (DBDpp) that specifically bind a target of interest. Nucleic acids encoding the DBDpp, and vectors and host cells containing the nucleic acids are also provided. Libraries of DBDpp, methods of producing and screening such libraries and the DBDpp identified from such libraries and screens are also encompassed. Methods of making and using the DBDpp are additionally provided. Such uses include, without limitation, affinity purification, and diagnostic and therapeutic applications.