Methods and reagents for analyzing protein-protein interfaces
AssigneeRevolution Medicines, Inc.
InventorsGregory L. VERDINE, M. James NICHOLS, Sharon A. TOWNSON, Uddhav Kumar SHIGDEL, Seung-Joo LEE, Dylan T. STILES, Neville J. ANTHONY
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Abstract
The present disclosure provides methods and reagents useful for analyzing protein-protein interfaces such as interfaces between a presenter protein (e.g., a member of the FKBP family, a member of the cyclophilin family, or PIN1) and a target protein. In some embodiments, the target and/or presenter proteins are intracellular proteins. In some embodiments, the target and/or presenter proteins are mammalian proteins.
Claims23
A method of covalently modulating KRAS, the method comprising contacting the KRAS with a presenter protein/compound complex, wherein
- the presenter protein is a member of the cyclophilin family, and
- the compound comprises
- a presenter protein binding moiety that binds to a cyclophilin protein,
- and a cross-linking group that reacts with an amino acid of KRAS to form a covalent bond between the compound and KRAS.
A method of identifying a compound capable of covalently binding to KRAS in the presence of a cyclophilin protein, said method comprising:
- (a) providing a sample comprising (i) a compound comprising a cyclophilin binding moiety and a cross-linking group;
- (ii) KRAS;
- and (iii) a cyclophilin protein;
- and
- (b) determining if said compound and KRAS form a covalent bond via the cross-linking group of said compound in said sample,
- wherein a compound is identified as covalently binding to KRAS in the presence of a cyclophilin protein if said compound and KRAS react in said sample.
A method of identifying a compound capable of selective and covalent binding to KRAS in the presence of a cyclophilin protein, said method comprising:
- (a) providing a first sample comprising (i) a compound comprising a cyclophilin binding moiety and a cross-linking group;
- (ii) KRAS;
- and (iii) a cyclophilin protein and a second sample comprising (i) the same compound comprising a cyclophilin binding moiety and a cross-linking group as in the first sample and (ii) KRAS as in the first sample;
- and
- (b) determining the extent to which said compound and KRAS react in said first sample as compared to said second sample,
- wherein a compound is identified as selectively covalently binding to KRAS in the presence of a cyclophilin protein if said compound and KRAS react in said first sample more than in said second sample.
A method of identifying compounds capable of modulating the biological activity of KRAS through covalent interaction, the method comprising:
- (a) providing the structure of a protein-protein interface in a complex comprising a cyclophilin protein and KRAS;
- and
- (b) determining the structure of compounds capable of binding at the interface, thereby identifying compounds capable of modulating the biological activity of KRAS.
A method of identifying compounds capable of modulating the biological activity of KRAS, the method comprising:
- (a) providing the structure of a protein-protein interface in a complex comprising a cyclophilin protein and KRAS;
- and
- (b) determining the structure of compounds capable of binding at the interface, thereby identifying compounds capable of modulating the biological activity of KRAS.
Dependencies are inferred from the claim text.