W0229

High Throughput Protein Structure Determination Using a Novel SAD Phasing Protocol. Qun Liu¹, Qingqiu Huang¹, Maikun Teng², Liwen Niu²and Quan Hao^{1, 1}MacCHESS, Wilson Synchrotron Lab, Cornell Univ., NY 14853, USA, Key Laboratory of Structural Biology, Chinese Academy of Science, Univ. of Science and Technology of China, Hefei, Anhui 230026, China.

A SAD phasing protocol for high throughput protein structure determination has been implemented and tested at MacCHESS. The method involves the coordination of three programs - SAPI, ABS and OASIS, which are developed at MacCHESS and distributed/supported by the CCP4.

SAPI is a direct-method based program and can be used to find a heavy atom substructure with single-wavelength anomalous scattering data, usually in several minutes. A built-in Karle-recycle process further refines the substructure.

The ABS program based on an algorithm proposed by Woolfson and Yao (1994, Acta Cryst. D50, 7-10) has been written to determine the absolute configuration (hand) of heavy atom sites by using anomalous scattering data. It also calculates a real-space figure of merit (FOM) that can be used to assess the quality of the solution for the anomalous scatter sites. Only the phases calculated from the correct hand are passed to the next phasing step.

OASIS is a well-established program used specifically to resolve the phase ambiguity intrinsic in SAD and SIR phasing.

To use the above three programs together with DM and Arp/Warp, available in the CCP4 Suite, it is feasible to solve a novel crystal structure in half an hour. Successful test has been performed with a SAD data set to 1.5 Å resolution collected for a new neurotoxin crystal at the MacCHESS F2 station. Three Cu sites were identified by using the SAPI program. After 30 minutes of computation on an ALPHA XP1000 processor running SAPI – ABS – OASIS – DM – Arp/Warp, 100 residues out of a total 124 could be automatically traced. The result has shown the applicability of integrating those programs together for high throughput structure determination that is essential for structural genomics and proteomics.