AB-INITIO STRUCTURE DETERMINATION AND REFINEMENT OF A SCORPION PROTEIN TOXIN

The structure of toxin II from the scorpion Androctonus australis Hect or has been determined ab initio by direct methods using SnB at 0.96 A ngstrom resolution. For the purpose of this structure redetermination, undertaken as a test of the minimal function and the SnB program, the identity and sequence of the protein was withheld from part of the re search team. A single solution obtained from 1 619 random atom trials was clearly revealed by the bimodal distribution of the final value of the minimal function associated with each individual trial. Five pept ide fragments were identified from a conservative analysis of the init ial E-map, and following several refinement cycles with X-PLOR, a mode l was built of the complete structure. At the end of the X-PLOR refine ment, the sequence was compared with the published sequence and 57 of the 64 residues had been correctly identified. Two errors in sequence resulted from side chains with similar size while the rest of the erro rs were a result of severe disorder or high thermal motion in the side chains. Given the amino-acid sequence, it is estimated that the initi al E-map could have produced a model containing 99% of all main-chain and 81% of side-chain atoms. The structure refinement was completed wi th PROFFT, including the contributions of protein H atoms, and converg ed at a residual of 0.158 for 30 609 data with F greater than or equal to 2 sigma(F) in the resolution range 8.0-0.964 Angstrom. The final m odel consisted of 518 non-H protein atoms (36 disordered), 407 H atoms , and 129 water molecules (43 with occupancies less than unity). This total of 647 non-H atoms represents the largest light-atom structure s olved to date.