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US 6,190,920 B1Granted
Method for detecting enzyme catalyzed cyclization
AssigneeThe Regents of the University of California
InventorsPaul A. Bartlett, Matthew T. Burger
Life of this patent
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Abstract
A method for detecting cyclization of acyclic compounds is disclosed. In particular, the invention relates to a method of screening for macrocyclic peptidase inhibitors, and is useful for screening a combinatorial library of compounds.
Claims20
A method for detecting enzyme-catalyzed cyclization of acyclic amino compounds, comprising;
- a) contacting a composition suspected of containing peptidase with an acyclic amino compound of Formula I:
- NH2—R1—X—R2—C(O)—Y
- wherein:
- C(O)Y represents a carboxylic acid or an ester or amide that can be acylated or hydrolyzed by said peptidase
- R1 and R2 each comprises one or more amino acid residues connected by covalent bonds and forming a structure that acts as a substrate of said peptidase, in which
- one of said amino acid residues in R1 is linked to a dye molecule or a resin, and
- one of said amino acid residues in R2 is linked to a resin or to a dye molecule, respectively, to provide a dye molecule attached on one side of X in said formula and a resin attached on the other side of X in said formula;
- and
- X is a group cleavable under reaction conditions that do not cleave an amide said group comprising an ester group, a disulfide group, a —CH(OH)—CH(OH)— group, or a carbonate group;
- b) contacting the product of step a) with an X-cleaving agent under said reaction conditions that do not cleave an amide;
- c) isolating the resin;
- and
- d) determining the presence or absence of the dye molecule on the isolated resin;
- wherein cyclization of said acyclic amino compound and presence of said peptidase is detected by retention of the dye molecule on the resin.
11 more elements
A method of discovering a peptidase inhibitor of Formula II from a collection of potential peptidase inhibitors, comprising:
- a) contacting a peptidase with an acyclic amino compound of Formula I:
- NH2—R1—X—R2—C(O)—Y
- wherein:
- C(O)Y represents a carboxylic acid or an ester or amide that can be acylated or hydrolyzed by said peptidase
- R1 and R2 each comprises one or more amino acid residues connected by covalent bonds, in which
- one of said amino acid residues in R1 is linked to a dye molecule or a resin, and
- one of said amino acid residues in R2 is linked to a resin or to a dye molecule, respectively, to provide a dye molecule attached on one side of X in said formula and a resin attached on the other side of X in said formula;
- and
- X is a group cleavable under reaction conditions that do not cleave an amide said group comprising an ester group, a disulfide group a —CH(OH)—CH(OH)— group, or a carbonate group;
- b) contacting the product of step a) with an X-cleaving agent under said reaction conditions that do not cleave an amide;
- c) isolating the resin;
- and
- d) determining the presence or absence of the dye molecule on the isolated resin;
- wherein ability of said compound of Formula I to act as an inhibitor of said peptidase is detected by retention of the dye molecule on the resin, thereby identifying as a peptidase inhibitor a compound of Formula II:
- where R1, R2, and X are as defined above.
12 more elements
Dependencies are inferred from the claim text.