Method and compositions for determining enzymatic activity and specificity of methyltransferases

Abstract

The invention provides crystalline O-methyltransferases and isolated non-native O-methyltransferases as well as sets of their structural coordinates. Also provided are methods of predicting the activity or substrate specificity of putative O-methyltransferases, methods of identifying potential substrates of O-methyltransferases, and methods of identifying potential inhibitors of methyltransferases.

1 . A method of predicting the activity and/or substrate specificity of a putative O-methyltransferase, said method comprising:
 comparing a three-dimensional representation of a known O-methyltransferase and the three-dimensional representation of said putative O-methyltransferase, wherein similarities and/or differences between the two representations are predictive of O-methyltransferase activity and/or substrate specificity.   
     
     
         2 . The method of  claim 1 , wherein the known O-methyltransferase is Chalcone O-methyltransferase (ChOMT) or isoflavone O-methyltransferase (IOMT). 
     
     
         3 . The method of  claim 2 , wherein the known O-methyltransferase has structural coordinates as set forth in Appendix A, Appendix B, Appendix C, Appendix D, PDB Accession No: 1FPQ, PDB Accession No: 1FPX, PDB Accession No: 1FP 1, or PDB Accession No: 1FP2. 
     
     
         4 . The method of  claim 1 , wherein the putative O-methyltransferase is a mutant of a known O-methyltransferase. 
     
     
         5 . A crystalline form of an O-methyltransferase having a set of structural coordinates as set forth in Appendix A, Appendix B, Appendix C, Appendix D, PDB Accession No: 1FPQ, PDB Accession No: 1FPX, PDB Accession No: 1FP1, or PDB Accession No: 1FP2. 
     
     
         6 . The crystalline O-methyltransferase of  claim 5 , wherein the O-methyltransferase is a Chalcone O-methyltransferase (ChOMT) or isoflavone O-methyltransferase(IOMT). 
     
     
         7 . A crystalline complex comprising an O-methyltransferase and an O-methyltransferase substrate. 
     
     
         8 . The crystalline complex of  claim 7 , wherein the O-methyltransferase is Chalcone O-methyltransferase (ChOMT) or isoflavone O-methyltransferase (IOMT). 
     
     
         9 . The crystalline complex of  claim 7 , wherein the O-methyltransferase is a non-native O-methyltransferase. 
     
     
         10 . The crystalline complex of  claim 7 , wherein the O-methyltransferase substrate is S-adenosyl-L-methionine (SAM). 
     
     
         11 . The crystalline complex of  claim 7 , wherein the complex has atomic coordinates as set forth in Appendix A, Appendix B, Appendix C, Appendix D, PDB Accession No: 1FPQ, PDB Accession No: 1FPX, PDB Accession No: 1FP1, or PDB Accession No: 1FP2. 
     
     
         12 . A method of identifying a potential substrate of an isoflavone O-methyltransferase, said method comprising:
 (a) defining an active site of said isoflavone O-methyltransferase based on a plurality of atomic coordinates of said isoflavone O-methyltransferase;   (b) identifying a potential substrate that fits the active site of (a); and   (c) contacting the isoflavone O-methyltransferase with the potential substrate and determining its activity thereon.   
     
     
         13 . (canceled) 
     
     
         14 . The method of  claim 12 , wherein the isoflavone O-methyltransferase is a mutant of a known isoflavone O-methyltransferase. 
     
     
         15 . (canceled) 
     
     
         16 . The method of  claim 12 , wherein the plurality of atomic coordinates are as set forth in Appendix B or Appendix D, or portions thereof. 
     
     
         17 . A method of identifying a potential inhibitor of an isoflavone O-methyltransferase, said method comprising:
 (a) contacting a potential compound that fits an active site based on a plurality of atomic coordinates of said isoflavone O-methyltransferase; and   (b) determining the ability of said compound to inhibit the activity of said isoflavone O-methyltransferase.   
     
     
         18 . (canceled) 
     
     
         19 . The method of  claim 17 , wherein the isoflavone O-methyltransferase is a mutant of a known isoflavone O-methyltransferase. 
     
     
         20 . (canceled) 
     
     
         21 . The method of  claim 17 , wherein the plurality of atomic coordinates are as set forth in Appendix B or Appendix D, or portions thereof. 
     
     
         22 .- 23 . (canceled)