Combinatorial libraries of monomer domains
OwnerJoost A. Kolkman, Willem P.C. Stemmer +1inventors
InventorsJoost A. Kolkman, Willem P.C. Stemmer, Sridhar Govindarajan
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Abstract
Methods for identifying discrete monomer domains and immuno-domains with a desired property are provided. Methods for generating multimers from two or more selected discrete monomer domains are also provided, along with methods for identifying multimers possessing a desired property. Presentation systems are also provided which present the discrete monomer and/or immuno-domains, selected monomer and/or immuno-domains, multimers and/or selected multimers to allow their selection. Compositions, libraries and cells that express one or more library member, along with kits and integrated systems, are also included in the present invention.
Claims112
A method for identifying a multimer that binds to a target molecule, the method comprising,
- providing a library of monomer domains;
- screening the library of monomer domains for affinity to a first target molecule;
- identifying at least one monomer domain that bind to at least one target molecule;
- linking the identified monomer domains to form a library of multimers, each multimer comprising at least two monomer domains;
- screening the library of multimers for the ability to bind to the first target molecule;
- and
- identifying a multimer that binds to the first target molecule.
A method for identifying a multimer that binds to at least one target molecule, the method comprising:
- providing a library of multimers, wherein each multimer comprises at least two monomer domains and each monomer domain exhibits a binding specificity for a target molecule;
- and
- screening the library of multimers for target molecule-binding multimers.
A library of multimers, wherein
- each multimer comprises at least two monomer domains connected by a linker;
- and
- each monomer domain exhibits a binding specificity for a target molecule.
48 The library of claim 47, wherein each monomer domain of the multimers is a non-naturally occurring monomer domain.
A polypeptide comprising at least two monomer domains separated by a heterologous linker, wherein each monomer domain specifically binds to a target molecule.
A method for identifying a multimer that binds to a target molecule, the method comprising,
- providing a library of immuno-domains;
- screening the library of immuno-domains for affinity to a first target molecule;
- identifying one or more (e.g., two or more) immuno-domains that bind to at least one target molecule;
- linking the identified monomer domain to form a library of multimers, each multimer comprising at least three immuno-domains (e.g., four or more, five or more, six or more, etc.);
- screening the library of multimers for the ability to bind to the first target molecule;
- and
- identifying a multimer that binds to the first target molecule.
A method of identifying hetero-immuno multimers that binds to a target molecule, the method comprising,
- providing a library of immuno-domains;
- screening the library of immuno-domains for affinity to a first target molecule;
- providing a library of monomer domains;
- screening the library of monomer domains for affinity to a first target molecule;
- identifying at least one immuno-domain that binds to at least one target molecule;
- identifying at least one monomer domain that binds to at least one target molecule;
- linking the identified immuno-domain with the identified monomer domains to form a library of multimers, each multimer comprising at least two domains;
- screening the library of multimers for the ability to bind to the first target molecule;
- and
- identifying a multimer that binds to the first target molecule.
A method for generating a library of chimeric monomer domains derived from human proteins, said method comprising:
- providing loop sequences corresponding to at least one loop from each of at least two different naturally occurring variants of a human protein, wherein the loop sequences are polynucleotide or polypeptide sequences;
- covalently combining loop sequences to generate a library of at least two different chimeric sequences, wherein each chimeric sequence encodes a chimeric monomer domain having at least two loops.
A method for identifying an LDL-receptor class A monomer domain that binds to a target molecule, the method comprising,
- providing a library of LDL-receptor class A monomer domains;
- screening the library of LDL-receptor class A monomer domains for affinity to a target molecule;
- and
- identifying an LDL-receptor class A monomer domain that binds to the target molecule.
A method for generating a library of chimeric LDL receptor A-domain monomers derived from human LDL receptor A-domains, said method comprising:
- providing loop sequences corresponding to at least one loop from each of two different naturally occurring variants of a human LDL receptor A-domains, wherein the loop sequences are polynucleotide or polypeptide sequences;
- covalently combining loop sequences to generate a library of chimeric monomer domain sequences, each chimeric sequence encoding a chimeric LDL receptor A-domain monomer having at least two loops;
- expressing the library of chimeric LDL receptor A-domain monomers using a display format selected from the group consisting of phage display, ribosome display, polysome display, and cell surface display;
- screening the expressed library of chimeric LDL receptor A-domain monomers for binding to a target molecule
- identifying a chimeric LDL receptor A-domain monomer domain that binds to the target molecule.
A method for identifying a multimer that binds to a target molecule, the method comprising:
- providing a library of monomer domains;
- screening the library of monomer domains for affinity to a first target molecule;
- identifying at least one monomer domain that bind to at least one target molecule;
- linking the identified at least one monomer domain to each member of the library of monomer domains to form a library of multimers;
- screening the library of multimers for the ability to bind to the first target molecule with an increased affinity;
- and
- identifying a multimer that binds to the first target molecule with an increased affinity.
A method for identifying a human chimeric monomer domain that binds to a target molecule, said method comprising:
- providing a sequence alignment of at least two naturally occurring human monomer domains from the same family of monomer domains;
- identifying amino acid residues in corresponding positions in the human monomer domain sequences that differ between the human monomer domains;
- generating a library of human chimeric monomer domains, wherein each human chimeric monomer domain sequence consists of amino acid residues that correspond in type and position to residues from two or more naturally occurring human monomer domains from the same family of monomer domains;
- screening the library of human chimeric monomer domains for binding to a target molecule;
- and
- identifying a human chimeric monomer domain that binds to a target molecule.
Dependencies are inferred from the claim text.