CRYOGENIC X-RAY CRYSTAL-STRUCTURE ANALYSIS FOR THE COMPLEX OF SCYTALONE DEHYDRATASE OF A RICE BLAST FUNGUS AND ITS TIGHT-BINDING INHIBITOR,CARPROPAMID - THE STRUCTURAL BASIS OF TIGHT-BINDING INHIBITION

Scytalone dehydratase is a member of the group of enzymes involved in fungal melanin biosynthesis in a phytopathogenic fungus, Pyricularia o ryzae, which causes rice blast disease. Carpropamid [( yl)ethyl]-1-eth yl-3-methylcyclopropanecarboxamide] is a tight-binding inhibitor of th e enzyme. To clarify the structural basis for tight-binding inhibition , the crystal structure of the enzyme complexed with carpropamid was a nalyzed using diffraction data collected at 100 K. The structural mode l was refined to a crystallographic R-factor of 0.180 against reflecti ons up to a resolution of 2.1 Angstrom. Carpropamid was bound in a hyd rophobic cavity of the enzyme. Three types of interactions appeared to contribute to the binding, (i) A hydrogen bond was formed between a c hloride atom in the dichloromethylethylcyclopropane ring of carpropami d and Asn-131 of the enzyme. (ii) The (chlorophenyl)ethyl group of car propamid built strong contacts with Val-75, and this group further for med a cluster of aromatic rings together with four aromatic residues i n the enzyme (Tyr-50, Phe-53, Phe-158, and Phe-162), (iii) Two hydrati on water molecules bound to the carboxamide group of carpropamid, and they were further hydrogen-bonded to Tyr-30, Tyr-50, His-85, and His-1 10. As a result of interactions between carpropamid and the phenylalan ine residues (Phe-158 and Phe-162) in the C-terminal region of the enz yme, the C-terminal region completely covered the inhibitor, ensuring its localization in the cavity.