CRYSTAL STRUCTURES OF HIGHLY SPECIFIC ASPERGILLUS RIBOTOXINS, RESTRICTOCIN AND MITOGILLIN. Xiaojing Yang¹, Sergio Martinez², Janet Smith² and Keith Moffat¹, ¹Department of Biochemistry and Molecular Biology, The University of Chicago, Chicago, IL 60637, USA and ²Department of Biological Sciences, Purdue University, West Lafayatte, IN 47907, USA

Two crystal structures in the Aspergillus ribotoxin family, restrictocin and mitogillin, are reported. The Aspergillus ribotoxins are a group of ribonucleases that specifically cleave a single phosphodiester bond in a highly conserved region of eukaryotic 28S ribosomal RNAs and thereby inhibit protein synthesis. The crystal structure of restrictocin is determined by single isomorphous replacement and anomalous scattering techniques and refined to 1.7 resolution using synchrotron Laue diffraction data. The crystal structure of mitogillin is determined by the difference Fourier method benefited by the high isomorphism between restrictocin and mitogillin crystals. The overall structures of restrictocin and mitogillin are identical except one residue where Ser25 of restrictocin is replaced by Asp25 of mitogillin. The crystal structure reveals a structural core in which a 5-stranded b-sheet is packed against a 3-turn a-helix, which can be well aligned with that of ribonuclease T1. Large peripheral loops near the active site construct a concave surface for substrate RNA binding. A lysine-rich loop is suggested to be responsible for the high substrate specificity by a docking model derived from the restrictocin structure and an NMR structure of a 29-mer RNA substrate analog. A large loop domain is also indicated to be involved in the cell entry activity of the Aspergillus ribotoxins.