European Molecular Biology Laboratory

Irma Querques

Orsolya Barabas

Cecilia Ines Zuliani

The present invention relates to improved transposase polypeptides having increased solubility. The enzyme of the invention was developed based on the Sleeping Beauty (SB) transposase. The invention provides further nucleic acids, vectors and recombinant cells encoding or containing the improved transposase, as well as a transposase system. Furthermore provided are medical and non-medical uses of the transposase of the invention for gene delivery. The invention is in particular useful as a tool for gene delivery in genetically modified cell based therapeutic approaches for treating various diseases.

Transfection method using a sleeping beauty transposase variant with increased solubility that transfects a target cell when used in protein form

This disclosure provides improved reagents and methods for inserting a transgene into the genome of a living host cell. Sleeping beauty (SB) transposase is used in protein form rather than as a vector. This has been made possible by using rational mutagenesis in a particular region of the SB crystal structure to develop variant SB transposase protein that is more soluble. The changes increase solubility while maintaining transposase activity, thereby adapting the variant transposase for the purpose of promoting recombinant integration of a transgene into a target cell when used in protein form rather than as a polynucleotide vector. The modified transposase is highly soluble in electroporation buffer and thermostable during storage. When introduced into a host cell, it promotes integration of a transgene into the genome of the cell in a dose-dependent manner. It is degraded within 48 hours, thereby rapidly clearing transposase activity from the host cell.

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Zurich, Switzerland

Irma Querques