Dual-space fragment extension using SAD data

       Extensive case studies have been carried out. Results have been summarized in References.

Flow chart	Azurin	Lysozyme	Xylanase	Staged iteration results of the left samples

References:

[1] D.Q. Yao, S. Huang, J.W. Wang, Y.X. Gu, C.D. Zheng & H.F. Fan, N. Watanabe & I. Tanaka, Acta Cryst. D62, 883-890. (2006)
[2] N. Watanabe, Y. Kitago, I. Tanaka, J.W. Wang, Y.X. Gu, C.D. Zheng & H.F. Fan, Acta Cryst. D61, 1533-1540. (2005)

2004

I. Auto-tuning the error term in direct-method SAD phasing

       The probability formula of direct-method SAD phasing used in the program OASIS contains an error term, which is related to the lack-of-closure error. Previously in the error term, there is a constant to be determined empirically for each particular case. It has been found that improper choice of the constant led often to the failure of phase derivation. The problem is resolved by introducing a modified error term and a method of automatically tuning the associated scaling factor.

Electron density maps obtained by a default run of the modified OASIS followed by a default run of DM (Cowtan, K.).

Cu-SAD	S-SAD	Se-SAD	Br-SAD

II. Direct-method SAD phasing with reciprocal-space fragment extension

       Structure fragments obtained from automatic model building based on direct-method SAD phasing are fed back to the direct-method probability formula to strengthen the phasing power and to benefit automation.

Flowchart	Xylanase synchrotron S-SAD data

Rusticyanin synchrotron Cu-SAD data		Thaumatin Cr-Ka S-SAD data

References:

[1] J.W. Wang, J.R. Chen, Y.X. Gu, C.D. Zheng, F. Jiang and H.F. Fan, Acta Cryst. D60, 1987-1990. (2004)
[2] J.W. Wang, J.R. Chen, Y.X. Gu, C.D. Zheng, and H.F. Fan, Acta Cryst. D60, 1991-1996. (2004)

2003

SAD phasing, why direct methods?

        SAD phasing is becoming popular in solving protein structures. The essential point to success is to break the phase ambiguity intrinsic in SAD data. Mostly this is done using density-modification techniques. However, density modification does not directly make choice between the two possible SAD phases for each reflection. The use of density-modification techniques alone leads often not to the best results. Two kinds of phase information enable discrimination of phase doublets from SAD data. One is from the heavy atoms (anomalous scatterers), while the other from the phase relationship of direct methods. The former can be expressed by the Sim distribution, while the latter can be expressed by the Cochran distribution. Usually only the Sim distribution is used to yield initial phases for the subsequent density modification. We use instead the product of the Sim and the Cochran distributions to produce initial phases, which have been proved much better than that from only the Sim distribution. Density modification based on such phases has been proved capable of yielding much better results than that based on only the Sim distribution.

Comparison of SAD phasing with and without direct methods

Phase ambiguity intrinsic in SAD	Phase information available in SAD	Two different kinds of initial SAD phases	Phase probability disributions

Cumulative phase errors	HMT SET7/9	Rusticyanin	Azurin

Reference:

[1] J.W. Wang, J.R. Chen, Y.X. Gu, C.D. Zheng, F. Jiang, H.F. Fan, T.C. Terwilliger and Q. Hao, Acta Cryst. D60, 1244-1253. (2004)

	2002:         Conventional MIR phases of the known protein Rpe have been significantly improved by the direct-method procedure DMIR Y.X. Gu, W.R. Chang, T. Jiang, C.D. Zheng & H.F. Fan, Acta Cryst. A58, 547-551 (2002)
	2001:         The Fourier map from conventional MAD phasing of 4-wavelength anomalous diffraction data was improved by direct-method phasing of only the 2-wavelength subset of data Fan Hai-fu, lecture at the Euro Conference on Phasing Biological Macromolecules, Martina Franca, Italy, June 2001 Y.X. Gu, F. Jiang, B.D. Sha & H.F. Fan, Z. Krist. 217, 710-714 (2002)
	2001:         Direct-method aided MAD phasing --- the first example of improving MAD phases by direct methods Y.X. Gu, Y.D. Liu, Q. Hao. S.E. Ealick & H.F. Fan, Acta Cryst. D57, 250-253 (2001)
OASIS	2000:         The CCP4 supported program for direct-method phasing of One-wavelength Anomalous Scattering or Single Isomorphous Substitution (replacement) Data Hao, Q., Gu, Y.X., Zheng, C.D. & Fan, H.F., J Appl. Cryst. 33, 980-981 (2000)
	1999:         The first example of breaking the protein SIR phase ambiguity when replacing atoms are in centric arrangement Liu Yu-dong, Gu Yuan-xin, Zheng Chao-de, Q. Hao and Fan Hai-fu, Acta Cryst. D55, 846-848 (1999)
	1999:         Comparing the result of breaking OAS-phase ambiguity using direct methods and that without using direct methods Liu Yu-dong, I. Harvey, Gu Yuan-xin, Zheng Chao-de, He Yi-zong, Fan Hai-fu, S. S. Hasnain, and Q. Hao, Acta Cryst. D55, 1620-1622 (1999)
	1997:         The first example of solving an originally unknown protein by direct-method phasing of the 2Å OAS data H. F. Fan, Q. Hao, I. Harvey, S. S. Hasnain, Y. D. Liu, Y. X. Gu, C. D. Zheng & H. Ke, in "Direct Methods for Solving Macromolecular Structures" Ed. by S. Fortier, Kluwer Academic Publishes, The Netherlands, 1998, pp. 479-485.
	1996:         Direct-method phasing and solvent flattening treatment of the 1.9Å OAS data of the protein azurin led to an easily traceable Fourier map Zheng Xiao-feng, Fan Hai-fu, Q. Hao, F.E. Dodd & S.S. Hasnian, Acta Cryst. D52, 937-941 (1996)
	1995:         Test with the OAS data from the protein core streptavidin, which was originally solved using 3-wavelength MAD data Sha Bing-dong, Liu Shen-ping, Gu Yuan-xin, Fan Hai-fu, Hengming Ke, Yao Jia-xing & M. M. Woolfson, Acta Cryst. D51, 342-346 (1995)
	1990:         The first traceable Fourier map from direct-method phasing of the experimental 2Å OAS data of the protein aPP Fan Hai-fu, Hao Quan, Gu Yuan-xin, Qian Jin-zi, Zheng Chao-de & Hengming Ke, Acta Cryst. A46, 935-939 (1990)
	1988:         The first successful test on direct-method phasing of the 3Å experimental SIR data from the native Bence Jones protein Rhe and its Au-derivative Collaborative work by H.F. Fan and B.C. Wang, 1988 (see M.M. Woolfson and Fan, Hai-fu, Physical and Non-physical Methods of Solving Crystal Structures, Cambridge University Press, 1995. Pp. 164-166.)
	1985:         The first successful test on direct-mothod phasing of the 2Å experimental OAS data from the protein aPP Fan Hai-fu & Gu Yuan-xin, Acta Cryst. A41, 280-284 (1985)
	1965:         It was proposed that the phase ambiguity due to single-wavelength anomalous diffration (SAD) or single isomorphous replacement (SIR) could be resolved by direct methods. Fan Hai-fu, Chinese Phys. 1429-1435 (1965) see also Fan Hai-fu, Acta Phys. Sin. 21 1114-1118 (1965). (In Chinese)

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