CRYSTAL-STRUCTURE OF AN ACIDIC NEUROTOXIN FROM SCORPION BUTHUS-MARTENSII KARSCH AT 1.85 ANGSTROM RESOLUTION

The crystal structure of an acidic scorpion neurotoxin, BmK M8, purifi ed from Chinese scorpion Buthus martensii Karsch (BmK), has been deter mined by the molecular replacement method. It is the first structure o f an acidic sc-scorpion neurotoxin reported so far. The crystals adopt a symmetry of space group P2(1) and contain one molecule per asymmetr ic unit. The structure has been refined to an R factor of 18.1% using reflection data in the range of 8 to 1.85 Angstrom resolution, with st andard deviations from ideal geometry of 0.017 Angstrom and 2.43 degre es for bond length and angle, respectively. The 12 residues at the C t erminus with unknown sequence were determined by crystallographic refi nement. The refined model shows that the structural core, consisting o f a motif beta alpha beta beta, is similar to that of toxin II from An droctonus australis Hector (AaH II) or Variant 3 from Centruroides scu lpturatus Ewing (CsE V3). The three conformationally variable loops pr otruding from this structural core are different from that of AaH II, and especially from that of CsE V3. Compared with the most potent and basic alpha-toxin AaH II, the BmK M8 is a relatively inactive toxin (1 100 times less active than AaH II) with an unusually low isoelectric p oint (pI 5.3). Sequence alignment of the two toxins shows a difference of 26 residues (40.6%). Among them four basic or neutral residues in AaH II, namely Val10, Lys28, Val55 and Gly59, are changed to acidic gl utamate in BmK M8. The residues Glu10, Glu28 and Glu55 of BmK M8 are l ocated on a surface (Face B), opposite the ''conserved hydrophobic sur face'' (Face A). The latter is a functionally important area proposed by Fontecilla-Camps et al. Our observations suggest that in addition t o Face A, Face B may also be involved in the biological activity of sc orpion toxins. The structure of BmK M8 shows an evident conformational change of the alpha-amino group at the N terminus and a deorganizatio n of Arg2 caused by the mutation D53A. These structural changes may al so be responsible for the weak toxicity of BmK M8. In association with the information from chemical modifications, a multisite binding mode for toxin-receptor interaction and three ''toxic regions'' in relevan ce to the binding process, including Face A, Face B and Site C, are pr oposed. Face A, mainly consisting of Tyr5, 35, 47, the alpha-amino gro up, Arg2 and Asp3, may be more essential for the binding. Face B, main ly comprising conserved residues Tyr14, 21, Lys28 and Va155, may contr ibute to the high efficacy of the binding process and substitutions by acidic residues in this area could strongly weaken the toxic activity . Site C, formed by Lys58 and Arg62 at the C terminus and Arg41 and Ty r42 from loop 38-44, may be involved in binding site specificity. (C) 1996 Academic Press Limited.