Engineered heterodimeric protein domains
First Claim
1. A multidomain protein comprising at least first and second nonidentical engineered domains, each of the first and second engineered domains containing a protein-protein interaction interface comprising amino acid sequence segments derived from two or more naturally occurring homologous parent domains, thereby conferring on said first and second engineered domains assembly specificities distinct from assembly specificities of the parent domains, wherein the first and second engineered domains form heterodimers with one another preferentially over forming homodimers.
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Abstract
The present invention provides an engineered multidomain protein including at least two nonidentical engineered domains, each of which contains a protein-protein interaction interface containing amino acid sequence segments derived from two or more existing homologous parent domains, thereby conferring on the engineered domains assembly specificities distinct from assembly specificities of the parent domains. In particular, the engineered domains form heterodimers with one another preferentially over forming homodimers. Methods of designing and using the engineered proteins are also included.
148 Citations
35 Claims
- 1. A multidomain protein comprising at least first and second nonidentical engineered domains, each of the first and second engineered domains containing a protein-protein interaction interface comprising amino acid sequence segments derived from two or more naturally occurring homologous parent domains, thereby conferring on said first and second engineered domains assembly specificities distinct from assembly specificities of the parent domains, wherein the first and second engineered domains form heterodimers with one another preferentially over forming homodimers.
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19. A multidomain protein comprising at least first and second nonidentical engineered domains that meet at an interface, the interface of each of the first and second engineered domains comprising at least two amino acid sequence segments, each derived from a different naturally-occurring homologous parent domain, thereby conferring a assembly specificity distinct from the assembly specificity of the parent domains, wherein the first and second engineered domains form heterodimers with one another preferentially over forming homodimers.
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20. A multidomain protein comprising at least first and second nonidentical engineered domains that meet at an interface, wherein (1) the first and second engineered domains are derived from two or more naturally-occurring homologous parent domains, (2) the interface from the first engineered domain comprises at least one amino acid sequence segment interacting with an amino acid sequence segment on the interface of the second engineered domain derived from the same parent domain, and (3) the first and second engineered domains form heterodimers with one another preferentially over forming homodimers.
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21. A multimeric protein comprising
(i) a domain with an amino acid sequence derived from two or more homologous parent domains; -
(ii) an interaction surface on said domain that mediates multimerization and that comprises amino acids derived from more than one of said parent domains; and
wherein the specificity of multimerization is enhanced by the presence of amino acids from different parent domains. - View Dependent Claims (22)
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- 23. An engineered immunoglobulin domain containing a protein-protein interaction interface comprising amino acids from two or more parent immunoglobulin domains such that the protein-protein interaction interface confers on the engineered immunoglobulin domain assembly specificities that are distinct from assembly specificities of the parent immunoglobulin domains, wherein the engineered immunoglobulin domain is not an antibody variable domain.
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25. An engineered immunoglobulin superfamily constant domain containing a protein-protein interaction interface comprising amino acids from two or more parent immunoglobulin domains such that the protein-protein interaction interface confers on the engineered immunoglobulin domain interaction properties that are distinct from interaction properties of the parent immunoglobulin domains.
- 26. A multidomain protein comprising an engineered domain that comprises a protein-protein interaction interface, said domain being homologous to a family of naturally-occurring domains, said interface comprising amino acids that are found in corresponding sequence positions in two or more said naturally-occurring domains, said amino acids not all being found at corresponding sequence positions in any single member of said family of naturally-occurring domains.
- 28. A nucleic acid encoding a multidomain protein comprising at least first and second nonidentical engineered domains, each of the first and second engineered domains containing protein-protein interaction interface comprising amino acid sequence segments derived from two or more naturally occurring homologous parent domains, thereby conferring assembly specificities of said first and second engineered domains distinct from assembly specificities of the parent domains, wherein (1) the first and second engineered domains form heterodimers with one another preferentially over forming homodimers, and (2) the first and second engineered domains are not antibody variable domains.
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31. A method of designing a multidomain protein with domains that preferentially heterodimerize, the method comprising the steps of:
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(a) selecting a first polypeptide, a second polypeptide, a third polypeptide and a fourth polypeptide, wherein the first and third polypeptides dimerize with each other, but not with the second or fourth polypeptide, and wherein said second and fourth polypeptides dimerize with each other, (b) composing an amino acid sequence of a first domain from the first and the second polypeptides comprising at least one assembly element from the first polypeptide, and (c) composing an amino acid sequence of a second domain from the third and fourth polypeptides comprising at least one assembly element from the third polypeptide, such that the assembly elements from the first and third polypeptides assemble with each other, thereby promoting heterodimerization of the first and second domains. - View Dependent Claims (32, 33, 34, 35)
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Specification