Molecules comprising a calcineurin-like binding pocket and encoded data storage medium capable of graphically displaying them

Abstract

The present invention relates to crystallized molecules and molecular complexes which comprise the active site binding pocket or the FKBP12/FK506 binding pocket of calcineurin or close structural homologues to either binding pocket. This invention also relates to a data storage material encoded with the corresponding structure coordinates of those crystallized molecules or molecular complexes. Such data storage material is capable of displaying such molecules and molecular complexes as a graphical three-dimensional representation on a computer screen. In addition, this invention relates to methods of using the structure coordinates of those molecules or molecular complexes to solve the structure of homologous proteins. This invention also relates to methods of using the structure coordinates to screen and design compounds that bind to calcineurin or homologues thereof.

1 . A crystallized molecule or molecular complex comprising a binding pocket defined by structure coordinates of CnA amino acids 90, 91, 92, 118, 120, 121, 122, 150, 151, 156, 160, 199, 232, 253, 254, 256, 281, 282, 283, 284, 306, 311, 312, and 317 according to  FIG. 1 , or a homologue of said molecule or molecular complex wherein said homologue comprises a binding pocket that has a root mean square deviation from the backbone atoms of said amino acids of not more than 1.5 Å.  
   
   
       2 . The crystallized molecule or molecular complex according to  claim 1 , wherein said binding pocket is defined by structure coordinates of CnA amino acids 90, 91, 92, 118, 120, 121, 122, 150, 151, 156, 160, 199, 281, 282, 283, 306, 311, 232, and 254, according to  FIG. 1 , or a homologue of said molecule or molecular complex, wherein said homologue comprises a binding pocket that has a root mean square deviation from the backbone atoms of said amino acids of not more than 1.5 Å.  
   
   
       3 . A crystallized molecule or molecular complex comprising a binding pocket defined by structure coordinates of CnA amino acids 122, 124, 159, 160, 310, 312, 313, 314, 339, 341, 343, 344, 345, 347, 351, 352, 353, 354, 355, 356, 359, 360, and 363; and CnB amino acids 49, 50, 114, 115, 118, 119, 121, 122, 123, 123, 157, 158, 159, 161, and 162 according to  FIG. 1 , or a homologue of said molecule or molecular complex, wherein said homologue comprises a binding pocket that has a root mean square deviation from the backbone atoms of said amino acids of not more than 1.5 Å.  
   
   
       4 . The crystallized molecule or molecular complex according to  claim 1 , further comprising a second binding pocket defined by CnA amino acids 122, 124, 159, 160, 310, 312, 313, 314, 339, 341, 343, 344, 345, 347, 351, 352, 353, 354, 355, 356, 359, 360, and 363; and CnB amino acids 49, 50, 114, 115, 118, 119, 121, 122, 123, 124, 157, 158, 159, 161, and 162; according to  FIG. 1 , or a homologue of said molecule or molecular complex, wherein said homologue comprises a second binding pocket that has a root mean square deviation from the backbone atoms of said amino acids of not more than 1.5 Å.  
   
   
       5 . The crystallized molecule or molecular complex according to  claim 4 , wherein said molecule or molecular complex is defined by the set of structure coordinates according to  FIG. 1 , or a homologue thereof, wherein said homologue has a root mean square deviation from the backbone atoms of said amino acids of not more than 1.5 Å.  
   
   
       6 . The crystallized molecule or molecular complex according to  claim 4 , wherein said molecule or molecular complex comprises amino acids 17-392 of CnA, CnB, FKBP12 and FK506.  
   
   
       7 - 15 . (canceled)  
   
   
       16 . A method for preparing a CnA/CnB/FKBP12/FK506 crystal comprising the steps of: 
 a. forming a molecular complex between FKBP12, FK506, calcineurin A and calcineurin B, wherein the calcineurin A lacks a calmodulin binding domain and an autoinhibitory domain; and    b. crystallizing the digested complex.    
   
   
       17 . The method according to  claim 16 , wherein the calmodulin binding domain and the autoinhibitory domain of said calcineurin A are removed by proteolytic digestion with a protease selected from clostripain, trypsin, endoproteinase Lys-C, endoproteinase Asp-N, endoproteinase Glu-C, elastase, enterokinase, restriction protease Factor Xa, thermolysin, Il-1 beta converting enzyme or HIV-1 protease.  
   
   
       18 . The method according to  claim 17 , wherein the protease is clostripain and the calcineurin A subunit in the crystallized complex has a molecular weight of about 42 kDa.