Simulated-1g1c - Revision history

Karsten at 13:14, 24 March 2020

2020-03-24T13:14:47Z

← Older revision		Revision as of 15:14, 24 March 2020
Line 575:		Line 575:
	but it traces only about 62 residues. The density looks somewhat reasonable, though.		but it traces only about 62 residues. The density looks somewhat reasonable, though.

	The files [~~ftp~~://{{SERVERNAME}}/pub/xds-datared/1g1c/xds-simulated-1g1c-I.mtz xds-simulated-1g1c-I.mtz] and [~~ftp~~://{{SERVERNAME}}/pub/xds-datared/1g1c/xds-simulated-1g1c-F.mtz xds-simulated-1g1c-F.mtz] are available.		The files [https://{{SERVERNAME}}/pub/xds-datared/1g1c/xds-simulated-1g1c-I.mtz xds-simulated-1g1c-I.mtz] and [https://{{SERVERNAME}}/pub/xds-datared/1g1c/xds-simulated-1g1c-F.mtz xds-simulated-1g1c-F.mtz] are available.

	I refined against 1g1c.pdb:		I refined against 1g1c.pdb:
Line 623:		Line 623:
	showing that the anomalous completeness, and even the quality of the anomalous signal, can indeed be increased. I doubt, however, that going to three or more frames would improve things even more.		showing that the anomalous completeness, and even the quality of the anomalous signal, can indeed be increased. I doubt, however, that going to three or more frames would improve things even more.

	The MTZ files are at [~~ftp~~://{{SERVERNAME}}/pub/xds-datared/1g1c/xds-simulated-1g1c-F-2frames.mtz] and [~~ftp~~://{{SERVERNAME}}/pub/xds-datared/1g1c/xds-simulated-1g1c-I-2frames.mtz], respectively. They were of course obtained with XDSCONV.INP:		The MTZ files are at [https://{{SERVERNAME}}/pub/xds-datared/1g1c/xds-simulated-1g1c-F-2frames.mtz] and [https://{{SERVERNAME}}/pub/xds-datared/1g1c/xds-simulated-1g1c-I-2frames.mtz], respectively. They were of course obtained with XDSCONV.INP:
	INPUT_FILE=temp.ahkl		INPUT_FILE=temp.ahkl
	OUTPUT_FILE=temp.hkl CCP4_I		OUTPUT_FILE=temp.hkl CCP4_I
Line 638:		Line 638:
	</pre>		</pre>

	Using the default (see above) phenix.refine job, I obtain against the [~~ftp~~://{{SERVERNAME}}/pub/xds-datared/1g1c/xds-simulated-1g1c-F-2frames.mtz MTZ file with amplitudes]:		Using the default (see above) phenix.refine job, I obtain against the [https://{{SERVERNAME}}/pub/xds-datared/1g1c/xds-simulated-1g1c-F-2frames.mtz MTZ file with amplitudes]:
	Start R-work = 0.3434, R-free = 0.3540		Start R-work = 0.3434, R-free = 0.3540
	Final R-work = 0.2209, R-free = 0.2479		Final R-work = 0.2209, R-free = 0.2479
	and against the [~~ftp~~://{{SERVERNAME}}/pub/xds-datared/1g1c/xds-simulated-1g1c-I-2frames.mtz MTZ file with intensities]		and against the [https://{{SERVERNAME}}/pub/xds-datared/1g1c/xds-simulated-1g1c-I-2frames.mtz MTZ file with intensities]
	Start R-work = 0.3492, R-free = 0.3606		Start R-work = 0.3492, R-free = 0.3606
	Final R-work = 0.2244, R-free = 0.2504		Final R-work = 0.2244, R-free = 0.2504
Line 754:		Line 754:
	total 165799 42025 43003 97.7% 11.7% 12.3% 162399 10.07 13.5% 14.8% 17% 0.908 16219		total 165799 42025 43003 97.7% 11.7% 12.3% 162399 10.07 13.5% 14.8% 17% 0.908 16219

	Using these data (stored in [~~ftp~~://{{SERVERNAME}}/pub/xds-datared/1g1c/xscale.oldversion]), I was finally able to solve the structure (see screenshot below) - SHELXE traced 160 out of 198 residues. All files produced by SHELXE are in [~~ftp~~://{{SERVERNAME}}/pub/xds-datared/1g1c/shelx].		Using these data (stored in [https://{{SERVERNAME}}/pub/xds-datared/1g1c/xscale.oldversion]), I was finally able to solve the structure (see screenshot below) - SHELXE traced 160 out of 198 residues. All files produced by SHELXE are in [https://{{SERVERNAME}}/pub/xds-datared/1g1c/shelx].

	[[File:1g1c-shelxe.png]]		[[File:1g1c-shelxe.png]]

Karsten at 16:33, 19 December 2019

2019-12-19T16:33:59Z

← Older revision		Revision as of 18:33, 19 December 2019
Line 575:		Line 575:
	but it traces only about 62 residues. The density looks somewhat reasonable, though.		but it traces only about 62 residues. The density looks somewhat reasonable, though.

	The files [ftp://~~turn5.biologie.uni-konstanz.de~~/pub/xds-datared/1g1c/xds-simulated-1g1c-I.mtz xds-simulated-1g1c-I.mtz] and [ftp://~~turn5.biologie.uni-konstanz.de~~/pub/xds-datared/1g1c/xds-simulated-1g1c-F.mtz xds-simulated-1g1c-F.mtz] are available.		The files [ftp://{{SERVERNAME}}/pub/xds-datared/1g1c/xds-simulated-1g1c-I.mtz xds-simulated-1g1c-I.mtz] and [ftp://{{SERVERNAME}}/pub/xds-datared/1g1c/xds-simulated-1g1c-F.mtz xds-simulated-1g1c-F.mtz] are available.

	I refined against 1g1c.pdb:		I refined against 1g1c.pdb:
Line 623:		Line 623:
	showing that the anomalous completeness, and even the quality of the anomalous signal, can indeed be increased. I doubt, however, that going to three or more frames would improve things even more.		showing that the anomalous completeness, and even the quality of the anomalous signal, can indeed be increased. I doubt, however, that going to three or more frames would improve things even more.

	The MTZ files are at [ftp://~~turn5.biologie.uni-konstanz.de~~/pub/xds-datared/1g1c/xds-simulated-1g1c-F-2frames.mtz] and [ftp://~~turn5.biologie.uni-konstanz.de~~/pub/xds-datared/1g1c/xds-simulated-1g1c-I-2frames.mtz], respectively. They were of course obtained with XDSCONV.INP:		The MTZ files are at [ftp://{{SERVERNAME}}/pub/xds-datared/1g1c/xds-simulated-1g1c-F-2frames.mtz] and [ftp://{{SERVERNAME}}/pub/xds-datared/1g1c/xds-simulated-1g1c-I-2frames.mtz], respectively. They were of course obtained with XDSCONV.INP:
	INPUT_FILE=temp.ahkl		INPUT_FILE=temp.ahkl
	OUTPUT_FILE=temp.hkl CCP4_I		OUTPUT_FILE=temp.hkl CCP4_I
Line 638:		Line 638:
	</pre>		</pre>

	Using the default (see above) phenix.refine job, I obtain against the [ftp://~~turn5.biologie.uni-konstanz.de~~/pub/xds-datared/1g1c/xds-simulated-1g1c-F-2frames.mtz MTZ file with amplitudes]:		Using the default (see above) phenix.refine job, I obtain against the [ftp://{{SERVERNAME}}/pub/xds-datared/1g1c/xds-simulated-1g1c-F-2frames.mtz MTZ file with amplitudes]:
	Start R-work = 0.3434, R-free = 0.3540		Start R-work = 0.3434, R-free = 0.3540
	Final R-work = 0.2209, R-free = 0.2479		Final R-work = 0.2209, R-free = 0.2479
	and against the [ftp://~~turn5.biologie.uni-konstanz.de~~/pub/xds-datared/1g1c/xds-simulated-1g1c-I-2frames.mtz MTZ file with intensities]		and against the [ftp://{{SERVERNAME}}/pub/xds-datared/1g1c/xds-simulated-1g1c-I-2frames.mtz MTZ file with intensities]
	Start R-work = 0.3492, R-free = 0.3606		Start R-work = 0.3492, R-free = 0.3606
	Final R-work = 0.2244, R-free = 0.2504		Final R-work = 0.2244, R-free = 0.2504
Line 754:		Line 754:
	total 165799 42025 43003 97.7% 11.7% 12.3% 162399 10.07 13.5% 14.8% 17% 0.908 16219		total 165799 42025 43003 97.7% 11.7% 12.3% 162399 10.07 13.5% 14.8% 17% 0.908 16219

	Using these data (stored in [ftp://~~turn5.biologie.uni-konstanz.de~~/pub/xds-datared/1g1c/xscale.oldversion]), I was finally able to solve the structure (see screenshot below) - SHELXE traced 160 out of 198 residues. All files produced by SHELXE are in [ftp://~~turn5.biologie.uni-konstanz.de~~/pub/xds-datared/1g1c/shelx].		Using these data (stored in [ftp://{{SERVERNAME}}/pub/xds-datared/1g1c/xscale.oldversion]), I was finally able to solve the structure (see screenshot below) - SHELXE traced 160 out of 198 residues. All files produced by SHELXE are in [ftp://{{SERVERNAME}}/pub/xds-datared/1g1c/shelx].

	[[File:1g1c-shelxe.png]]		[[File:1g1c-shelxe.png]]

Kay: /* Finally solving the structure */

2011-04-10T20:52:33Z

Finally solving the structure

← Older revision		Revision as of 22:52, 10 April 2011
Line 724:		Line 724:
	=== Finally solving the structure ===		=== Finally solving the structure ===

	After thinking about the most likely way that James Holton used to produce the simulated data, I hypothesized that within each frame, the radiation damage is most likely constant, and that there is a jump in radiation damage from frame 1 to 2. Unfortunately for this scenario, the scaling algorithm in CORRECT and XSCALE was changed for the version of Dec-2010, such that it produces best results when the changes are smooth. Therefore, I tried the penultimate version of XSCALE - and indeed that gives significantly better results:		After thinking about the most likely way that James Holton used to produce the simulated data, I hypothesized that within each frame, the radiation damage is most likely constant, and that there is a jump in radiation damage from frame 1 to 2. Unfortunately for this scenario, the scaling algorithm in CORRECT and XSCALE was changed for the version of Dec-2010, such that it produces best results when the changes are smooth. Therefore, I tried the penultimate version (May-2010) of XSCALE - and indeed that gives significantly better results:

	NOTE: Friedel pairs are treated as different reflections.		NOTE: Friedel pairs are treated as different reflections.
Line 755:		Line 755:

	Using these data (stored in [ftp://turn5.biologie.uni-konstanz.de/pub/xds-datared/1g1c/xscale.oldversion]), I was finally able to solve the structure (see screenshot below) - SHELXE traced 160 out of 198 residues. All files produced by SHELXE are in [ftp://turn5.biologie.uni-konstanz.de/pub/xds-datared/1g1c/shelx].		Using these data (stored in [ftp://turn5.biologie.uni-konstanz.de/pub/xds-datared/1g1c/xscale.oldversion]), I was finally able to solve the structure (see screenshot below) - SHELXE traced 160 out of 198 residues. All files produced by SHELXE are in [ftp://turn5.biologie.uni-konstanz.de/pub/xds-datared/1g1c/shelx].

	[[File:1g1c-shelxe.png]]		[[File:1g1c-shelxe.png]]
	It is worth mentioning that James Holton confirmed that my hypothesis is true; he also ~~mentions~~ that this approach is a good approximation for a multi-pass data collection.
	However, generally the smooth scaling gives better results than the previous method of assigning the same scale factor to all reflections of a frame; in particular, it correctly treats those reflections near the border of two frames. This example shows that it is important to		It is worth mentioning that James Holton confirmed that my hypothesis is true; he also says that this approach is a good approximation for a multi-pass data collection.

			However, generally (i.e. for real data) the smooth scaling (which also applies to absorption correction and detector modulation) gives better results than the previous method of assigning the same scale factor to all reflections of a frame; in particular, it correctly treats those reflections near the border of two frames.

			Phenix.refine against these data gives:
			Start R-work = 0.3449, R-free = 0.3560
			Final R-work = 0.2194, R-free = 0.2469
			which is only 0.15%/0.10% better in R-work/R-free than the previous best result (see above).

			This example shows that it is important to
	* have the best data available if a structure is difficult to solve		* have the best data available if a structure is difficult to solve
	* know the options (programs, algorithms)		* know the options (programs, algorithms)
	* know as much as possible about the experiment		* know as much as possible about the experiment

Kay: /* Solving the structure */

2011-04-10T20:33:05Z

Solving the structure

← Older revision		Revision as of 22:33, 10 April 2011
Line 530:		Line 530:
	but there does not appear a "magic bullet" that would produce much better data than with the quick bootstrap approach.		but there does not appear a "magic bullet" that would produce much better data than with the quick bootstrap approach.

	== ~~Solving~~ the structure ==		== Trying to solve the structure ==

	First, we repeat xscale after inserting FRIEDEL'S_LAW=FALSE into XSCALE.INP . This gives us		First, we repeat xscale after inserting FRIEDEL'S_LAW=FALSE into XSCALE.INP . This gives us

Kay: /* Why this is difficult to solve with SAD phasing */

2011-04-10T20:31:24Z

Why this is difficult to solve with SAD phasing

← Older revision		Revision as of 22:31, 10 April 2011
Line 720:		Line 720:

	so the strongest peak corresponds to the translation of molecules (0,0.5,0) but the origin peak is at 1/2 of that size, which appears significant.		so the strongest peak corresponds to the translation of molecules (0,0.5,0) but the origin peak is at 1/2 of that size, which appears significant.


			=== Finally solving the structure ===

			After thinking about the most likely way that James Holton used to produce the simulated data, I hypothesized that within each frame, the radiation damage is most likely constant, and that there is a jump in radiation damage from frame 1 to 2. Unfortunately for this scenario, the scaling algorithm in CORRECT and XSCALE was changed for the version of Dec-2010, such that it produces best results when the changes are smooth. Therefore, I tried the penultimate version of XSCALE - and indeed that gives significantly better results:

			NOTE: Friedel pairs are treated as different reflections.

			SUBSET OF INTENSITY DATA WITH SIGNAL/NOISE >= -3.0 AS FUNCTION OF RESOLUTION
			RESOLUTION NUMBER OF REFLECTIONS COMPLETENESS R-FACTOR R-FACTOR COMPARED I/SIGMA R-meas Rmrgd-F Anomal SigAno Nano
			LIMIT OBSERVED UNIQUE POSSIBLE OF DATA observed expected Corr

			8.05 1922 467 476 98.1% 4.0% 5.8% 1888 22.37 4.5% 2.5% 84% 1.952 142
			5.69 3494 864 882 98.0% 4.7% 6.0% 3429 20.85 5.4% 3.2% 77% 1.707 297
			4.65 4480 1111 1136 97.8% 5.1% 5.9% 4395 21.13 5.8% 3.3% 68% 1.518 406
			4.03 5197 1325 1357 97.6% 5.3% 6.0% 5101 20.57 6.1% 3.8% 48% 1.280 499
			3.60 5915 1500 1533 97.8% 6.0% 6.3% 5803 19.99 6.9% 4.1% 41% 1.169 572
			3.29 6601 1657 1694 97.8% 6.5% 6.5% 6476 19.42 7.5% 4.6% 27% 1.066 634
			3.04 7080 1789 1830 97.8% 7.6% 7.2% 6948 17.50 8.7% 5.4% 23% 1.037 693
			2.85 7682 1945 1979 98.3% 8.8% 9.0% 7528 14.75 10.1% 7.0% 15% 0.935 750
			2.68 8099 2062 2100 98.2% 11.0% 11.1% 7933 12.81 12.7% 9.1% 13% 0.881 795
			2.55 8351 2155 2201 97.9% 13.3% 13.7% 8178 11.16 15.4% 11.0% 12% 0.872 836
			2.43 9195 2327 2376 97.9% 16.5% 17.2% 9003 9.49 19.0% 15.1% 8% 0.838 904
			2.32 9495 2377 2428 97.9% 19.8% 20.3% 9304 8.62 22.7% 17.3% 4% 0.818 934
			2.23 9936 2498 2551 97.9% 20.8% 21.7% 9751 8.30 23.9% 17.5% 4% 0.830 987
			2.15 10217 2577 2622 98.3% 23.3% 24.0% 9990 7.74 26.7% 19.2% 4% 0.814 998
			2.08 10710 2704 2766 97.8% 27.1% 28.6% 10506 6.82 31.1% 23.5% 5% 0.812 1071
			2.01 10899 2777 2839 97.8% 28.1% 29.2% 10648 6.46 32.3% 25.0% 6% 0.813 1059
			1.95 11361 2878 2937 98.0% 34.4% 35.5% 11134 5.55 39.5% 30.3% 3% 0.780 1136
			1.90 11639 2941 3000 98.0% 40.5% 41.5% 11403 4.88 46.6% 35.9% 0% 0.787 1163
			1.85 12020 3068 3123 98.2% 52.2% 55.1% 11752 3.79 60.0% 47.4% 6% 0.775 1195
			1.80 11506 3003 3173 94.6% 60.8% 64.8% 11229 3.23 70.1% 58.8% 0% 0.765 1148
			total 165799 42025 43003 97.7% 11.7% 12.3% 162399 10.07 13.5% 14.8% 17% 0.908 16219

			Using these data (stored in [ftp://turn5.biologie.uni-konstanz.de/pub/xds-datared/1g1c/xscale.oldversion]), I was finally able to solve the structure (see screenshot below) - SHELXE traced 160 out of 198 residues. All files produced by SHELXE are in [ftp://turn5.biologie.uni-konstanz.de/pub/xds-datared/1g1c/shelx].
			[[File:1g1c-shelxe.png]]
			It is worth mentioning that James Holton confirmed that my hypothesis is true; he also mentions that this approach is a good approximation for a multi-pass data collection.
			However, generally the smooth scaling gives better results than the previous method of assigning the same scale factor to all reflections of a frame; in particular, it correctly treats those reflections near the border of two frames. This example shows that it is important to
			* have the best data available if a structure is difficult to solve
			* know the options (programs, algorithms)
			* know as much as possible about the experiment

Kay: /* Why this is difficult to solve with SAD phasing */

2011-04-05T14:46:15Z

Why this is difficult to solve with SAD phasing

← Older revision		Revision as of 16:46, 5 April 2011
Line 675:		Line 675:
	"This crystal form contains two molecules in the asymmetric unit. They are related by a noncrystallographic two-fold axis, parallel to the crystallographic b axis, located at X = 0.25 and Z = 0.23. This arrangement results in a peak in the native Patterson map at U = 0.5, V = 0, W = 0.47 of peak height 26 σ (42% of the origin peak)."		"This crystal form contains two molecules in the asymmetric unit. They are related by a noncrystallographic two-fold axis, parallel to the crystallographic b axis, located at X = 0.25 and Z = 0.23. This arrangement results in a peak in the native Patterson map at U = 0.5, V = 0, W = 0.47 of peak height 26 σ (42% of the origin peak)."

	Unfortunately, the arrangement of substructure sites has translational symmetry, and may be related to a centrosymmetric arrangement. Indeed, the original structure was solved using molecular replacement.		Unfortunately, the arrangement of substructure sites has (pseudo-)translational symmetry, and may be related to a centrosymmetric arrangement. Indeed, the original structure was solved using molecular replacement.

	Using the four sites as given by SHELXE (and default parameters otherwise), I obtained from the [http://cci.lbl.gov/cctbx/phase_o_phrenia.html cctbx - Phase-O-Phrenia server] the following		Using the four sites as given by SHELXE (and default parameters otherwise), I obtained from the [http://cci.lbl.gov/cctbx/phase_o_phrenia.html cctbx - Phase-O-Phrenia server] the following
Line 719:		Line 719:
	...		...

	so the strongest peak corresponds to the translation of molecules (0,0.5,0) but the origin peak is at 1/2 of that size, which appears significant. ~~(I cannot interpret the plot of relative peak heights, though <big>☹</big>)~~		so the strongest peak corresponds to the translation of molecules (0,0.5,0) but the origin peak is at 1/2 of that size, which appears significant.

Kay at 11:50, 5 April 2011

2011-04-05T11:50:36Z

← Older revision		Revision as of 13:50, 5 April 2011
Line 586:		Line 586:
	== Notes ==		== Notes ==

	=== Towards better completeness: using the first two frames ===		=== Towards better completeness: using the first two frames instead of only the first ===

	We might want better (anomalous) completeness than what is given by only the very first frame of each dataset. To this end, we change in the XDS.INP part of our script :		We might want better (anomalous) completeness than what is given by only the very first frame of each dataset. To this end, we change in the XDS.INP part of our script :
Line 645:		Line 645:
	Final R-work = 0.2244, R-free = 0.2504		Final R-work = 0.2244, R-free = 0.2504

	so: better R-free is obtained from better data.		so: '''better R-free is obtained from better data.'''

	The statistics from SHELXD and SHELXE don't look better - they were already quite good with a single frame per dataset. The statistics printed by SHELXE (for the correct hand) are:		The statistics from SHELXD and SHELXE don't look better - they were already quite good with a single frame per dataset. The statistics printed by SHELXE (for the correct hand) are:
Line 669:		Line 669:
	6 0.5158 0.4788 0.0406 4.7 5/1.45 3/4.94 4/7.85 1/21.38 5/23.30		6 0.5158 0.4788 0.0406 4.7 5/1.45 3/4.94 4/7.85 1/21.38 5/23.30

	Using ~~these~~ four sites (and ~~standard~~ parameters otherwise, I obtained from the [http://cci.lbl.gov/cctbx/phase_o_phrenia.html cctbx - Phase-O-Phrenia server] the following		=== Why this is difficult to solve with SAD phasing ===

			In the original publication ("Structural evidence for a possible role of reversible disulphide bridge formation in the elasticity of the muscle protein titin" Mayans, O., Wuerges, J., Canela, S., Gautel, M., Wilmanns, M. (2001) Structure 9: 331-340 ) we read:

			"This crystal form contains two molecules in the asymmetric unit. They are related by a noncrystallographic two-fold axis, parallel to the crystallographic b axis, located at X = 0.25 and Z = 0.23. This arrangement results in a peak in the native Patterson map at U = 0.5, V = 0, W = 0.47 of peak height 26 σ (42% of the origin peak)."

			Unfortunately, the arrangement of substructure sites has translational symmetry, and may be related to a centrosymmetric arrangement. Indeed, the original structure was solved using molecular replacement.

			Using the four sites as given by SHELXE (and default parameters otherwise), I obtained from the [http://cci.lbl.gov/cctbx/phase_o_phrenia.html cctbx - Phase-O-Phrenia server] the following
	Plot of relative peak heights:		Plot of relative peak heights:

Line 712:		Line 720:

	so the strongest peak corresponds to the translation of molecules (0,0.5,0) but the origin peak is at 1/2 of that size, which appears significant. (I cannot interpret the plot of relative peak heights, though <big>☹</big>)		so the strongest peak corresponds to the translation of molecules (0,0.5,0) but the origin peak is at 1/2 of that size, which appears significant. (I cannot interpret the plot of relative peak heights, though <big>☹</big>)

	~~=== Why this is difficult to solve with SAD phasing ===~~

	In the original publication ("Structural evidence for a possible role of reversible disulphide bridge formation in the elasticity of the muscle protein titin" Mayans, O., Wuerges, J., Canela, S., Gautel, M., Wilmanns, M. (2001) Structure 9: 331-340 ) we read:

	"This crystal form contains two molecules in the asymmetric unit. They are related by a noncrystallographic two-fold axis, parallel to the crystallographic b axis, located at X = 0.25 and Z = 0.23. This arrangement results in a peak in the native Patterson map at U = 0.5, V = 0, W = 0.47 of peak height 26 σ (42% of the origin peak)."

	Unfortunately this translates into a almost centrosymmetric arrangement of substructure sites (check with [http://cci.lbl.gov/cctbx/phase_o_phrenia.html]). Indeed, the original structure was solved using molecular replacement.

Kay: /* Towards better completeness: using the first two frames */

2011-04-05T10:44:09Z

Towards better completeness: using the first two frames

@@ Line 637: / Line 637: @@
 EOF
 </pre>
 === Why this is difficult to solve with SAD phasing ===

Kay: /* Towards better completeness: using the first two frames */

2011-03-22T16:05:54Z

Towards better completeness: using the first two frames

← Older revision		Revision as of 18:05, 22 March 2011
Line 588:		Line 588:
	=== Towards better completeness: using the first two frames ===		=== Towards better completeness: using the first two frames ===

	We might want better (anomalous) completeness than what is given by only the very first frame of each dataset. To this end, we change in XDS.INP :		We might want better (anomalous) completeness than what is given by only the very first frame of each dataset. To this end, we change in the XDS.INP part of our script :
	DATA_RANGE=1 2		DATA_RANGE=1 2
	~~and~~ in XSCALE.INP ~~we insert~~		then run the script which reduces the 100 datasets. When this has finished, we insert in XSCALE.INP
	NBATCH=2		NBATCH=2
	after each INPUT_FILE line. The reason for this is that by default, XSCALE establishes scalefactors every ~~DELPHI (default:~~ 5) degrees, but here we want scalefactors for every frame, because the radiation damage is so strong. This gives:		after each INPUT_FILE line (this can be easily done using <pre> awk '{print $0;print "NBATCH=2"}' XSCALE.INP > x </pre>). The reason for this is that by default, XSCALE establishes scalefactors every 5 degrees, but here we want scalefactors for every frame, because the radiation damage is so strong. This gives:
	NOTE: Friedel pairs are treated as different reflections.		NOTE: Friedel pairs are treated as different reflections.

Line 621:		Line 621:
	total 165827 42032 43003 97.7% 12.8% 13.3% 162426 8.79 14.7% 15.7% 15% 0.881 16225		total 165827 42032 43003 97.7% 12.8% 13.3% 162426 8.79 14.7% 15.7% 15% 0.881 16225

	showing that the anomalous completeness, and even the quality of the anomalous signal, can indeed be increased. I doubt, however, that going to three or more frames would improve things even more.		showing that the anomalous completeness, and even the quality of the anomalous signal, can indeed be increased. I doubt, however, that going to three or more frames would improve things even more.

			The MTZ files are at [ftp://turn5.biologie.uni-konstanz.de/pub/xds-datared/1g1c/xds-simulated-1g1c-F-2frames.mtz] and [ftp://turn5.biologie.uni-konstanz.de/pub/xds-datared/1g1c/xds-simulated-1g1c-I-2frames.mtz], respectively. They were of course obtained with XDSCONV.INP:
			INPUT_FILE=temp.ahkl
			OUTPUT_FILE=temp.hkl CCP4_I
			for the intensities, and
			INPUT_FILE=temp.ahkl
			OUTPUT_FILE=temp.hkl CCP4
			for the amplitudes. In both cases, after xdsconv we have to run
			<pre>
			f2mtz HKLOUT temp.mtz<F2MTZ.INP
			cad HKLIN1 temp.mtz HKLOUT output_file_name.mtz<<EOF
			LABIN FILE 1 ALL
			END
			EOF
			</pre>

	=== Why this is difficult to solve with SAD phasing ===		=== Why this is difficult to solve with SAD phasing ===

Kay: /* Solving the structure */

2011-03-22T15:55:00Z

Solving the structure

← Older revision		Revision as of 17:55, 22 March 2011
Line 583:		Line 583:
	Final R-work = 0.2170, R-free = 0.2596		Final R-work = 0.2170, R-free = 0.2596
	which appears reasonable.		which appears reasonable.

			== Notes ==

			=== Towards better completeness: using the first two frames ===

			We might want better (anomalous) completeness than what is given by only the very first frame of each dataset. To this end, we change in XDS.INP :
			DATA_RANGE=1 2
			and in XSCALE.INP we insert
			NBATCH=2
			after each INPUT_FILE line. The reason for this is that by default, XSCALE establishes scalefactors every DELPHI (default: 5) degrees, but here we want scalefactors for every frame, because the radiation damage is so strong. This gives:
			NOTE: Friedel pairs are treated as different reflections.

			SUBSET OF INTENSITY DATA WITH SIGNAL/NOISE >= -3.0 AS FUNCTION OF RESOLUTION
			RESOLUTION NUMBER OF REFLECTIONS COMPLETENESS R-FACTOR R-FACTOR COMPARED I/SIGMA R-meas Rmrgd-F Anomal SigAno Nano
			LIMIT OBSERVED UNIQUE POSSIBLE OF DATA observed expected Corr

			8.05 1922 467 476 98.1% 4.2% 6.6% 1888 20.04 4.8% 2.8% 84% 1.887 142
			5.69 3494 864 882 98.0% 4.5% 6.8% 3429 18.67 5.2% 3.1% 83% 1.635 297
			4.65 4480 1111 1136 97.8% 5.3% 6.7% 4395 18.89 6.1% 3.5% 66% 1.347 406
			4.03 5197 1325 1357 97.6% 6.2% 6.8% 5101 18.37 7.1% 4.3% 43% 1.156 499
			3.60 5916 1500 1533 97.8% 6.9% 7.1% 5804 17.83 8.0% 4.7% 36% 1.083 572
			3.29 6601 1657 1694 97.8% 7.6% 7.3% 6476 17.26 8.7% 4.9% 24% 1.029 634
			3.04 7081 1789 1830 97.8% 9.1% 8.0% 6949 15.50 10.4% 6.4% 17% 1.011 693
			2.85 7684 1946 1979 98.3% 10.9% 9.9% 7530 12.95 12.5% 8.1% 16% 0.950 751
			2.68 8101 2062 2100 98.2% 13.1% 12.1% 7935 11.18 15.0% 10.5% 10% 0.888 795
			2.55 8355 2156 2201 98.0% 15.2% 14.9% 8182 9.69 17.5% 12.3% 6% 0.867 837
			2.43 9195 2327 2376 97.9% 18.2% 18.6% 9003 8.20 20.8% 15.4% 6% 0.837 904
			2.32 9495 2377 2428 97.9% 21.3% 21.9% 9304 7.42 24.4% 18.4% 6% 0.800 934
			2.23 9939 2499 2551 98.0% 23.0% 23.3% 9753 7.13 26.4% 19.0% 4% 0.818 987
			2.15 10219 2577 2622 98.3% 25.4% 25.9% 9992 6.63 29.1% 20.6% 1% 0.797 998
			2.08 10712 2704 2766 97.8% 29.4% 30.8% 10508 5.80 33.8% 25.1% 4% 0.793 1071
			2.01 10900 2778 2839 97.9% 30.8% 31.2% 10649 5.50 35.3% 26.2% 4% 0.828 1060
			1.95 11361 2878 2937 98.0% 36.7% 38.2% 11134 4.71 42.1% 31.5% 1% 0.768 1136
			1.90 11641 2943 3000 98.1% 42.7% 45.1% 11405 4.12 49.1% 38.7% -1% 0.775 1165
			1.85 12028 3069 3123 98.3% 54.0% 60.4% 11760 3.19 62.1% 47.5% 5% 0.735 1196
			1.80 11506 3003 3173 94.6% 62.1% 70.6% 11229 2.72 71.6% 60.6% -2% 0.709 1148
			total 165827 42032 43003 97.7% 12.8% 13.3% 162426 8.79 14.7% 15.7% 15% 0.881 16225

			showing that the anomalous completeness, and even the quality of the anomalous signal, can indeed be increased. I doubt, however, that going to three or more frames would improve things even more.

			=== Why this is difficult to solve with SAD phasing ===

			In the original publication ("Structural evidence for a possible role of reversible disulphide bridge formation in the elasticity of the muscle protein titin" Mayans, O., Wuerges, J., Canela, S., Gautel, M., Wilmanns, M. (2001) Structure 9: 331-340 ) we read:

			"This crystal form contains two molecules in the asymmetric unit. They are related by a noncrystallographic two-fold axis, parallel to the crystallographic b axis, located at X = 0.25 and Z = 0.23. This arrangement results in a peak in the native Patterson map at U = 0.5, V = 0, W = 0.47 of peak height 26 σ (42% of the origin peak)."

			Unfortunately this translates into a almost centrosymmetric arrangement of substructure sites (check with [http://cci.lbl.gov/cctbx/phase_o_phrenia.html]). Indeed, the original structure was solved using molecular replacement.