The immunosuppressive agent mizoribine monophosphate forms a transition state analogue complex with inosine monophosphate dehydrogenase.
Mizoribine monophosphate (MZP) is the active metabolite of the immunosuppressive agent mizoribine and a potent inhibitor of IMP dehydrogenase (IMPDH). This enzyme catalyzes the oxidation of IMP to XMP with the concomitant reduction of NAD via a covalent intermediate at Cys319 (E-XMP). Surprisingly, mutational analysis indicates that MZP is a transition state analogue although its structure does not resemble that of the expected transition state. Here we report the X-ray crystal structure of the E.MZP complex at 2.0 A resolution that reveals a transition state-like structure and solves the mechanistic puzzle of the IMPDH reaction. The protein assumes a new conformation where a flap folds into the NAD site and MZP, Cys319, and a water molecule are arranged in a geometry resembling the transition state. The water appears to be activated by interactions with a conserved Arg418-Tyr419 dyad. Mutagenesis experiments confirm that this new closed conformation is required for the hydrolysis of E-XMP, but not for the reduction of NAD. The closed conformation provides a structural explanation for the differences in drug selectivity and catalytic efficiency of IMPDH isozymes.
Gan
Lu
L
Department of Biochemistry and Chemistry and the Rosenstiel Basic Medical Sciences Research Center, Brandeis University, Waltham, Massachusetts 02454, USA.
Seyedsayamdost
Mohammad R
MR
Shuto
Satoshi
S
Matsuda
Akira
A
Petsko
Gregory A
GA
Hedstrom
Lizbeth
L
D000818:Animals; D001667:Binding, Competitive; D002384:Catalysis; D006224:Cricetinae; D018360:Crystallography, X-Ray; D004791:Enzyme Inhibitors / Q000737:chemistry* / Q000378:metabolism; D007168:IMP Dehydrogenase / Q000037:antagonists & inhibitors / Q000737:chemistry* / Q000378:metabolism; D007166:Immunosuppressive Agents / Q000737:chemistry* / Q000378:metabolism; D008956:Models, Chemical; D011487:Protein Conformation; D012263:Ribonucleosides / Q000737:chemistry* / Q000378:metabolism; D016835:Tritrichomonas foetus / Q000201:enzymology; D014867:Water / Q000737:chemistry
D004791:Enzyme Inhibitors; D007166:Immunosuppressive Agents; D012263:Ribonucleosides; D014867:Water; C010052:mizoribine; D007168:IMP Dehydrogenase