US 2019/0153433 A1 · TRANSPOSITION-MEDIATED IDENTIFICATION OF SPECIFIC BINDING OR FUNCTIONAL PROTEINS

Abstract

The method disclosed herein describes a novel technology offering unparalleled efficiency, flexibility, utility and speed for the discovery and optimization of polypeptides having desired binding specificity and/or functionality, including antigen-binding molecules such as antibodies and fragments thereof, for desired functional and/or binding phenotypes. The novel method is based on transposable constructs and diverse DNA libraries cloned into transposable vectors and their transfection into host cells by concomitant transient expression of a functional transposase enzyme. This ensures an efficient, stable introduction of the transposon-based expression vectors into vertebrate host cells in one step, which can then be screened for a desired functional or binding phenotype of the expressed proteins, after which the relevant coding sequences for the expressed proteins, including antibodies and fragments thereof, can be identified by standard cloning and DNA sequencing techniques.

Claims23

Dependent claims

1-20. (canceled)

A method of obtaining a polypeptide having a desired binding specificity or functionality, comprising:

(i) generating a diverse collection of polynucleotides encoding polypeptides having different binding specificities or functionalities, wherein said polynucleotides each comprise a sequence encoding a polypeptide, said sequence disposed between first and second inverted terminal repeat sequences that are recognized by and functional with a least one transposase enzyme;
(ii) introducing the diverse collection of polynucleotides of (i) into host cells;
(iii) expressing at least one transposase enzyme functional with said inverted terminal repeat sequences in said host cells so that said diverse collection of polynucleotides is integrated into the host cell genomes to provide a host cell population that expresses said polypeptides having different binding specificities or functionalities;
(iv) screening said host cells to identify a host cell expressing a polypeptide having a desired binding specificity or functionality;
and
(v) isolating the polynucleotide sequence encoding said polypeptide identified in step (iv) from said host cell.

3 more elements

Dependencies are inferred from the claim text.

Transposition-mediated identification of specific binding or functional proteins

Life of this patent

Transposition-mediated identification of specific binding or functional proteins

Life of this patent

Abstract

Claims23