Abstract: Methods of generating fusion protein variants are provided that comprise introducing sequence diversity at the junction region or regions in the fusion and allows for the generation of variants having a desired activity. Examples include immunoglobulins comprising a domain or polypeptide inserted into, or replacing, a CDR. Also provided are polynucleotides encoding a fusion protein and comprising two or more RSSs, and compositions and host cells comprising same, as well as fusion proteins variants produced by the described methods.

Abstract: A receptor is provided having a heterologous binding site that activates, when bound, a signaling domain related to the TNF receptor superfamily. Methods/uses of the foregoing in whole cell biosensors are also provided. There is also provided a library comprising a plurality of unique biosensor cells for binding unknown binding substrates. Each unique biosensor is a host cell having a receptor which signals production of a positive selectable marker and/or a negative selectable marker in response to the receptor being bound. Also provided is a method of identifying biosensor cells from a library that is specifically activated by a target, involving (a) contacting the library with the target substrate under positive selection conditions; (b) contacting the library with a control substrate under negative selection conditions; and (c) identifying biosensor cells which survive (a) and (b) as biosensor cells which are specifically activated by the target.

Abstract: Methods of generating fusion protein variants are provided that comprise introducing sequence diversity at the junction region or regions in the fusion and allows for the generation of variants having a desired activity. Examples include immunoglobulins comprising a domain or polypeptide inserted into, or replacing, a CDR. Also provided are polynucleotides encoding a fusion protein and comprising two or more RSSs, and compositions and host cells comprising same, as well as fusion proteins variants produced by the described methods.

Abstract: Methods of generating sequence diversity in a protein, such as a ligand-binding protein, are provided. The methods comprise targeted introduction of two or more recombination signal sequences (RSSs) into the protein coding sequence and introduction of the modified protein coding sequence into a recombination-competent host cell, specifically a recombination-competent host cell that is capable of expressing at least RAG-1 and RAG-2, thereby allowing for recombination of the protein coding sequence and expression of variant proteins. Also provided are polynucleotides comprising a nucleic acid sequence encoding a target protein, such as a ligand-binding protein, and comprising two or more RSSs, and compositions and host cells comprising same.

Abstract: Methods of generating fusion protein variants are provided that comprise introducing sequence diversity at the junction region or regions in the fusion and allows for the generation of variants having a desired activity. Examples include immunoglobulins comprising a domain or polypeptide inserted into, or replacing, a CDR. Also provided are polynucleotides encoding a fusion protein and comprising two or more RSSs, and compositions and host cells comprising same, as well as fusion proteins variants produced by the described methods.

Abstract: Compositions and methods are disclosed for generating immunoglobulin structural diversity in vitro, and in particular, for reducing biases in V region and J segment gene utilization, and for generating immunoglobulin V-D-J recombination events in a manner that does not require D-J recombination to precede V-DJ recombination. Selection of advantageous combinations of immunoglobulin gene elements, including introduction of artificial diversity (D) segment genes and optimization of recombination signal sequence (RSS) efficiency, are disclosed.

Abstract: Compositions and methods are disclosed for generating immunoglobulin structural diversity in vitro, and in particular, for reducing biases in V region and J segment gene utilization, and for generating immunoglobulin V-D-J recombination events in a manner that does not require D-J recombination to precede V-DJ recombination. Selection of advantageous combinations of immunoglobulin gene elements, including introduction of artificial diversity (D) segment genes and optimization of recombination signal sequence (RSS) efficiency, are disclosed.