Enhanced Transposase Polypeptides for Mammalian Cell Engineering
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Solution Overview
Problem
Current transposon/transposase systems lack high-level activity in mammalian cells, limiting their application in genetic engineering for human/mammalian cell engineering.
Innovation Solution
Development of recombinant transposase polypeptides with enhanced activity in mammalian cells, specifically comprising sequences at least 90% identical to certain transposase sequences like hSB110 or hSB81, and featuring specific amino acid positions, which are used to facilitate the integration of genetic elements into mammalian cell genomes.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If conventional transposon/transposase systems are used, then the system structure is simple and well-established, but the transposase activity in mammalian cells is insufficient
Solution Approach 1:
The patent applies parameter changes by modifying specific amino acid residues in the transposase sequence (e.g., positions 136, 253, 255, 314) to enhance transposase activity in mammalian cells. These targeted sequence modifications change the biochemical parameters of the transposase enzyme, improving its functionality without completely redesigning the entire system.
Solution Approach 2:
The patent creates composite transposase constructs by combining modified transposase sequences with heterologous polypeptide sequences (such as purification tags or cell-penetrating peptides). This composite approach integrates the enhanced transposase activity with additional functional elements to achieve both high activity and practical utility in mammalian cell engineering.
2Productivity
If transposase activity is enhanced through sequence modification, then genetic engineering efficiency improves, but the deviation from natural transposase sequence increases
Solution Approach 1:
The patent applies local quality by making targeted modifications at specific positions (136, 253, 255, 314) within the transposase sequence rather than uniformly altering the entire sequence. This localized approach concentrates the functional improvements at critical sites while preserving the overall sequence structure and identity of the original transposase.
Solution Approach 2:
The patent implements partial action by introducing only the necessary minimal modifications (specific amino acid changes at key positions) required to achieve enhanced transposase activity in mammalian cells, rather than completely redesigning the transposase. This partial modification strategy achieves the desired productivity improvement while maintaining sequence stability.
Data Source
AI summary
Transposase polypeptides and polynucleotides are provided, which have a high activity in mammalian cells. Methods for engineering cells, such as chimeric antigen T-cells, with the transposes are also provided.