Crystal Structure of Aldehyde Dehydrogenase and Methods of Use Thereof

Abstract

The present disclosure provides a crystal structure of aldehyde dehydrogenase (ALDH) with a modulator of ALDH bound thereto. The present disclosure provides a computer readable medium comprising atomic coordinates for an ALDH polypeptide and a modulator bound to a site within the polypeptide. A method is also provided. In general terms, the method comprises computationally identifying a compound that binds to an ALDH polypeptide, using the atomic coordinates.

1 . A crystal comprising an aldehyde dehydrogenase (ALDH) polypeptide in crystalline form, wherein said crystal comprises a chemical entity bound to the active site of the ALDH polypeptide. 
     
     
         2 . The crystal of  claim 1 , wherein the ALDH polypeptide is an ALDH2 polypeptide, wherein the crystal is characterized with space group P2 1 , and has unit cell parameters of a=102 Å, b=177 Å, c=103 Å, bond angles of a=γ=90°, b=94.5°. 
     
     
         3 . The crystal of  claim 1 , wherein the ALDH polypeptide is an ALDH2 polypeptide that comprises a Glu at a position corresponding to amino acid 504 of SEQ ID NO:1, wherein the crystal is characterized with space group P2 1 , and has unit cell parameters of a=102 Å, b=177 Å, c=102 Å, bond angles of a=γ=90°, b=94.6° 
     
     
         4 . The crystal of  claim 1 , wherein the bound chemical entity is an agonist. 
     
     
         5 . The crystal of  claim 4 , wherein the agonist is N-(1,3-benzodioxo1-5-ylmethyl)-2,6-dichlorobenzamide. 
     
     
         6 . The crystal of  claim 1 , wherein the ALDH polypeptide has a length of about 500 amino acids. 
     
     
         7 . The crystal of  claim 1 , wherein the ALDH polypeptide comprises an amino acid sequence having at least about 80% amino acid sequence identity to amino acids 18-517 of the amino acid sequence set forth in SEQ ID NO:1, wherein the ALDH polypeptide comprises a Glu at a position corresponding to amino acid 504 of SEQ ID NO:1. 
     
     
         8 . The crystal of  claim 1 , wherein the ALDH polypeptide comprises an amino acid sequence having at least about 80% amino acid sequence identity to amino acids 18-517 of the amino acid sequence set forth in SEQ ID NO:1, wherein the ALDH polypeptide comprises a Lys at a position corresponding to amino acid 504 of SEQ ID NO:1. 
     
     
         9 . A composition comprising the crystal of  claim 1 . 
     
     
         10 . The composition of  claim 1 , wherein the crystal diffracts x-rays for a determination of structure coordinates to a resolution of between 1.5 Angstroms and 2.0 Angstroms. 
     
     
         11 . A method comprising:
 computationally identifying a compound that binds to an aldehyde dehydrogenase (ALDH) polypeptide using atomic coordinates for a complex comprising said ALDH polypeptide and a ligand bound to a ligand-binding site within the ALDH polypeptide.   
     
     
         12 . The method of  claim 11 , wherein said atomic coordinates are set forth in Table 1. 
     
     
         13 . The method of  claim 11 , wherein said atomic coordinates are set forth in Table 6. 
     
     
         14 . The method of  claim 11 , further comprising:
 testing said compound to determine if it modulates an enzymatic activity of said ALDH polypeptide.   
     
     
         15 . The method of  claim 11 , further comprising:
 testing said compound to determine if it modulates a substrate specificity of said ALDH polypeptide.   
     
     
         16 . The method of  claim 11 , wherein said computationally identifying employs a docking program that computationally tests known compounds for binding to said ALDH polypeptide. 
     
     
         17 . The method of  claim 11 , wherein said computationally identifying includes designing a compound that binds to said ALDH polypeptide. 
     
     
         18 . The method of  claim 17 , wherein said compound is designed from a known compound. 
     
     
         19 . A method comprising:
 a) receiving a set of atomic coordinates for a complex comprising an aldehyde dehydrogenase (ALDH) polypeptide and a ligand bound to a ligand-binding site within said ALDH polypeptide;   b) identifying a compound that binds to said ALDH polypeptide using said coordinates.   
     
     
         20 . A method of identifying a drug candidate compound for the treatment of a disorder, the method comprising:
 a) employing the three-dimensional structural coordinates of an aldehyde dehydrogenase (ALDH) polypeptide and determining the binding mode of a test compound within the catalytic site of the polypeptide;   b) selecting a test compound having the best fit with the ALDH catalytic site; and   c) assaying the ability of the test compound to modulate ALDH catalytic activity, wherein a test compound that modulates ALDH catalytic activity is considered a candidate agent for treating a disorder.   
     
     
         21 . The method of  claim 20 , wherein the test agent blocks access of a substrate to one or both of Cys 302 and Glu 268 of the active site, wherein the test agent reduces catalytic activity of the ALDH polypeptide, and wherein the test agent is considered a candidate agent for treating a disorder that would benefit from reducing ALDH activity. 
     
     
         22 . The method of  claim 21 , wherein the disorder is cancer, and wherein the test agent is considered a candidate agent for sensitizing a cancer cell to a cancer chemotherapeutic agent. 
     
     
         23 . The method of  claim 20 , wherein the test agent increases binding of a substrate to one or both of Cys 302 and Glu 268 of the active site, wherein the test agent increases catalytic activity of the ALDH polypeptide, and wherein the test agent is considered a candidate agent for treating a disorder that would benefit from increasing ALDH activity. 
     
     
         24 . The method of  claim 23 , wherein the disorder is a disorder resulting from a toxic level of an aldehyde, cataract, oral cancer, esophageal cancer, an upper digestive tract cancer, lung cancer, atopic dermatitis, radiation dermatitis, an acute or chronic ischemic or oxidative stress disease, nitroglycerin insensitivity, seizure, or a neurodegenerative disease. 
     
     
         25 . A computer-assisted method for identifying potential modulators of aldehyde dehydrogenase (ALDH), using a programmed computer comprising a processor, a data storage system, an input device, and an output device, the method comprising:
 a) inputting into the programmed computer through said input device data comprising the three-dimensional coordinates of a subset of the atoms generated from a complex of ALDH and an agonist or an antagonist bound at or near the active site of the ALDH, thereby generating a criteria data set;   b) comparing, using said processor, said criteria data set to a computer database of chemical structures stored in said computer data storage system;   c) selecting from said database, using computer methods, chemical structures having a portion that is structurally similar to said criteria data set; and   d) outputting to said output device the selected chemical structures having a portion similar to said criteria data set.   
     
     
         26 . A computer readable medium comprising:
 atomic coordinates for a complex comprising: i) an aldehyde dehydrogenase (ALDH) polypeptide; and ii) a ligand bound to a ligand-binding site in the ALDH polypeptide.   
     
     
         27 . The computer readable medium of  claim 26 , further comprising:
 programming for displaying a molecular model of said ALDH polypeptide.   
     
     
         28 . The computer readable medium of  claim 26 , further comprising:
 programming for identifying a compound that binds to said ALDH polypeptide.   
     
     
         29 . The computer readable medium of  claim 28 , further comprising:
 a database of structures of known test compounds.   
     
     
         30 . The computer readable medium of  claim 26 , wherein said atomic coordinates are set forth in Table 1 or in Table 6. 
     
     
         31 . A computer comprising the computer-readable medium of  claim 26 . 
     
     
         32 . A computer system comprising: a memory comprising X-ray crystallographic structure coordinates defining a ligand-binding site of a complex comprising an aldehyde dehydrogenase (ALDH) polypeptide with a ligand bound to a ligand-binding site within the ALDH polypeptide;
 and a processor in electrical communication with the memory; wherein the processor generates a molecular model having a three dimensional structure representative of at least a portion of said ALDH polypeptide-bound ligand complex.