Hello crystallography experts,
I think I may have a case of perfect pseudomerehedral twinning where P222
Laue
symmetry is actually P2 with twinning and aniostropic data masks intensity
based
twinning statistics. I would like some input to determine if my reasoning
makes sense.
I would like to perform a twin test using local intensity differences (J E
Padilla and T O Yeates, 2003) but I am not sure how to do this. I am a
little worried that using a twin law in a lower symmetry group than
appropriate will give the drop in Rfree/Rwork that I am seeing and the
crystal is not really twinned.
1. I have processed in P222 and P21. In each case I get good scaling
statistics using XDS (see below)
2. Systematic absences clearly point to a single 2(1)-fold screw axis
3. There appears to "almost" be two more 2(1) screw axis except that there
are some significant
violations in these absences leading me to believe that they are not
true srew axis.
4. Patterson map shows no peaks relating molecules by translation in P222
(1 molec/asu)
5. I have difficulty interpreting the patterson map in P2 (2molec/asu) (I
am new to this so could be my own fault)
7. Intensity statistics show no twinning but the data is very aniosotropic
suggesting that twinning can be masked. The cumulative distribuiton of z=
(I/<I>) deviates for centric reflections my a max of 0.104 and the curve
falls well below for all values of (Phenix triage).
8. Molecular replacement (Phaser) in P222 gives a single solution in
P212121. Every other choice of space group fails to give a solution.
9. Refinement in P212121 space group stalls around 34/31% Rfree/Rwork.
Althrough there are geometry problems to correct and water has not been
added.
10. Molecular replacement in P2 give single solution in P21
11. Refinement against the same model used above gives 36.5 33.8
Rfree/Rwork without twin law
and 32/30 Rfree/Rwork with twin law = h, -k, -l
I have shown scaling for P212121. Statistics are nearly the same for P222
scaling
except XDS throws up a warning that I have outliers in my systematic
absences.
P212121 20-1.7A
CELL 56.279 67.257 106.264
Res OBS UNIQUE POS COMP R(obs) R(exp) I/Sig Rmeas CC1/2
4.96 13696 1911 1989 96.1% 2.6% 2.7% 13681 61.96 2.8% 99.9* 23*
1.044 1400
3.56 24387 3163 3167 99.9% 2.5% 2.8% 24386 64.65 2.7% 99.9* 9
0.888 2672
2.93 22483 3969 3995 99.3% 3.2% 3.1% 22463 46.05 3.5% 99.9* 1
0.880 3292
2.54 20108 4685 4712 99.4% 4.2% 4.1% 20076 30.45 4.8% 99.8* 1
0.839 3710
2.28 20966 5221 5268 99.1% 5.5% 5.6% 20923 22.69 6.3% 99.7* -1
0.779 3932
2.08 21599 5702 5779 98.7% 7.3% 7.7% 21521 16.73 8.5% 99.4* 0
0.756 3932
1.93 21589 6216 6289 98.8% 11.1% 12.1% 21478 10.73 13.1% 98.5* 0
0.735 3755
1.80 19392 579 6709 98.1% 17.5% 19.7% 19089 6.32 21.3% 96.1* -3
0.695 2421
1.70 10889 5093 7126 71.5% 24.8% 27.6% 9561 3.63 31.7% 89.2* 2
0.750 350
total 175109 42539 45034 94.5% 4.0% 4.2% 173178 23.25 4.6% 99.9* 1
0.808 25464
0 21 0 3.203 0.6806E+03 0.3320E+02 20.50 3*
0 22 0 3.057 0.9423E+04 0.3288E+03 28.66 1
0 23 0 2.924 0.3205E+03 0.4586E+02 6.99 1*
0 24 0 2.802 0.1231E+03 0.4612E+02 2.67 1
0 25 0 2.690 0.2306E+02 0.4566E+02 0.51 1*
0 26 0 2.587 0.1540E+04 0.7494E+02 20.56 1
0 27 0 2.491 0.3212E+03 0.5579E+02 5.76 1*
0 28 0 2.402 0.2706E+04 0.1156E+03 23.42 1
0 29 0 2.319 0.3761E+02 0.4931E+02 0.76 1*
0 30 0 2.242 0.4053E+03 0.6758E+02 6.00 1
0 31 0 2.170 0.4495E+03 0.6956E+02 6.46 1*
0 32 0 2.102 0.3573E+04 0.1442E+03 24.78 1
0 33 0 2.038 0.5625E+02 0.7052E+02 0.80 1*
0 34 0 1.978 0.2167E+04 0.1090E+03 19.88 1
0 35 0 1.922 0.3802E+02 0.6082E+02 0.63 1*
0 36 0 1.868 0.1628E+04 0.9449E+02 17.23 1
3 0 0 18.760 0.6193E+01 0.5268E+01 1.18 2*
4 0 0 14.070 0.4882E+04 0.1672E+03 29.19 1
5 0 0 11.256 0.3065E+02 0.8641E+01 3.55 2*
7 0 0 8.040 0.1511E+03 0.1280E+02 11.81 2*
8 0 0 7.035 0.5131E+04 0.1255E+03 40.87 2
9 0 0 6.253 -0.2260E-01 0.1419E+02 0.00 2*
10 0 0 5.628 0.7113E+03 0.2502E+02 28.43 2
11 0 0 5.116 0.3705E+03 0.2093E+02 17.70 2*
13 0 0 4.329 0.4986E+03 0.2448E+02 20.37 2*
15 0 0 3.752 0.8513E+03 0.3547E+02 24.00 2*
16 0 0 3.517 0.7354E+03 0.3602E+02 20.42 2
17 0 0 3.311 0.2188E+03 0.3146E+02 6.95 2*
18 0 0 3.127 0.8876E+04 0.2186E+03 40.60 2
19 0 0 2.962 0.3268E+02 0.3828E+02 0.85 1*
20 0 0 2.814 0.5687E+03 0.4339E+02 13.11 1
21 0 0 2.680 0.9416E+01 0.3746E+02 0.25 1*
22 0 0 2.558 0.6604E+03 0.4963E+02 13.31 1
23 0 0 2.447 0.1710E+02 0.4819E+02 0.35 1*
24 0 0 2.345 0.6804E+03 0.6019E+02 11.30 1
25 0 0 2.251 -0.7221E+02 0.5214E+02 -1.38 1*
26 0 0 2.165 0.8755E+03 0.6379E+02 13.72 1
27 0 0 2.084 0.5401E+02 0.5433E+02 0.99 1*
28 0 0 2.010 0.4692E+03 0.5928E+02 7.92 1
29 0 0 1.941 0.1127E+03 0.5774E+02 1.95 1*
AVERAGE INTENSITY FOR 95 REFLECTIONS WHICH SHOULD
BE SYSTEMATICALLY ABSENT IS 11.3% OF MEAN INTENSITY
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 CC(1/2) Anomal SigAno NanoLIMIT
OBSERVED UNIQUE POSSIBLE OF DATA observed expected
Corr
4.96 13696 1911 1989 96.1% 2.6% 2.7%
13681 61.96 2.8% 99.9* 23*
1.044 1400
3.56 24387 3163 3167 99.9% 2.5% 2.8%
24386 64.65 2.7% 99.9* 9 0.888 2672
2.93 22483 3969 3995 99.3% 3.2% 3.1%
22463 46.05 3.5% 99.9* 1 0.880 3292
2.54 20108 4685 4712 99.4% 4.2% 4.1%
20076 30.45 4.8% 99.8* 1 0.839 3710
2.28 20966 5221 5268 99.1% 5.5% 5.6%
20923 22.69 6.3% 99.7* -1 0.779 3932
2.08 21599 5702 5779 98.7% 7.3% 7.7%
21521 16.73 8.5% 99.4* 0 0.756 3932
1.93 21589 6216 6289 98.8% 11.1% 12.1%
21478 10.73 13.1% 98.5* 0 0.735 3755
1.80 19392 6579 6709 98.1% 17.5% 19.7%
19089 6.32 21.3% 96.1* -3 0.695 2421
1.70 10889 5093 7126 71.5% 24.8% 27.6%
9561 3.63 31.7% 89.2* 2 0.750 350
total 175109 42539 45034 94.5% 4.0% 4.2%
173178 23.25 4.6% 99.9* 1 0.808 25464
Very low res data complete to ~90% (10-20A)
twinning analysis
Accentric
<I**2>/<I>**2 = 1.96 (untwinned 2.0, pefect twin 1.5)
<|L|> = 0.478 (0.5 expected, 0.375, perfect twin)
<L**2> = 0.314 (0.33 expected, perfect twin .20)
Wilson plot - estimated B factor = 10.6
1 molec /asu
1 off origin patterson peak Height: 4.127
Molecular replacement (Phaser) and Refinement (Phenix)
Soution only in P212121 when processed in P222
After refinement r_work = 0.2760 r_free = 0.3221 bonds = 0.011 angles
= 1.430
Waters not yet added
Still some work to be done: Only 88% in Favored regions and a helix to
be modeledin (10 out of ~ 400 residues)
--------------------------------------------------------------------------------------------------------------------------------
56.188 106.072 67.149 90 90.083 90
P2
REFLECTIONS OF TYPE H,0,0 0,K,0 0,0,L OR EXPECTED TO BE ABSENT (*)
--------------------------------------------------------------------
H K L RESOLUTION INTENSITY SIGMA INTENSITY/SIGMA #OBSERVED
0 0 4 16.787 0.7371E+03 0.1419E+02 51.93 4
0 0 5 13.430 -0.1886E+00 0.6700E+01 -0.03 3
0 0 6 11.191 0.8114E+03 0.1904E+02 42.62 3
0 0 7 9.593 0.1002E+02 0.8988E+01 1.11 3
0 0 8 8.394 0.1207E+04 0.3247E+02 37.18 2
0 0 9 7.461 0.2742E+03 0.1516E+02 18.09 2
0 0 10 6.715 0.2812E+04 0.5793E+02 48.54 3
0 0 11 6.104 0.9688E+01 0.1350E+02 0.72 3
0 0 12 5.596 0.5878E+04 0.1171E+03 50.21 3
0 0 13 5.165 0.1393E+02 0.1553E+02 0.90 3
0 0 14 4.796 0.4194E+03 0.2053E+02 20.43 3
0 0 17 3.950 0.1953E+03 0.2314E+02 8.44 3
0 0 18 3.730 0.3026E+04 0.6746E+02 44.86 3
0 0 19 3.534 0.4052E+03 0.2912E+02 13.92 3
0 0 20 3.357 0.3184E+04 0.7203E+02 44.21 3
0 0 22 3.052 0.9765E+04 0.3365E+03 29.02 1
0 0 23 2.920 0.3395E+03 0.4897E+02 6.93 1
0 0 24 2.798 0.1234E+03 0.4774E+02 2.58 1
0 0 25 2.686 0.1902E+02 0.4727E+02 0.40 1
0 0 26 2.583 0.1635E+04 0.7802E+02 20.95 1
0 0 27 2.487 0.3243E+03 0.5796E+02 5.60 1
0 0 28 2.398 0.2913E+04 0.1222E+03 23.83 1
0 0 29 2.315 0.3650E+02 0.5151E+02 0.71 1
0 0 30 2.238 0.4204E+03 0.6762E+02 6.22 1
0 0 31 2.166 0.4294E+03 0.6832E+02 6.29 1
0 0 32 2.098 0.3665E+04 0.1455E+03 25.18 1
0 0 33 2.035 0.4843E+02 0.6951E+02 0.70 1
0 0 34 1.975 0.2151E+04 0.1070E+03 20.10 1
0 0 35 1.919 0.3692E+02 0.5922E+02 0.62 1
0 0 36 1.865 0.1430E+04 0.8572E+02 16.69 1
0 6 0 17.679 0.2716E+04 0.6484E+02 41.88 2
0 7 0 15.153 0.5690E+01 0.5728E+01 0.99 2
0 9 0 11.786 -0.8183E+01 0.7247E+01 -1.13 2
0 10 0 10.607 0.7695E+03 0.2079E+02 37.01 2
0 11 0 9.643 -0.1919E+01 0.8897E+01 -0.22 2
0 12 0 8.839 0.1198E+04 0.3096E+02 38.70 2
0 13 0 8.159 0.9875E+01 0.1048E+02 0.94 2
0 15 0 7.071 0.2055E+02 0.1179E+02 1.74 2
0 17 0 6.240 0.1361E+01 0.1252E+02 0.11 2
0 18 0 5.893 0.5356E+04 0.1290E+03 41.54 2
0 19 0 5.583 0.4309E+02 0.1458E+02 2.95 2
0 20 0 5.304 0.5713E+04 0.1377E+03 41.49 2
0 21 0 5.051 0.3815E+02 0.1559E+02 2.45 2
0 22 0 4.821 0.1963E+04 0.5152E+02 38.10 2
0 23 0 4.612 -0.4939E+02 0.1813E+02 -2.72 2
0 24 0 4.420 0.6401E+03 0.2774E+02 23.08 2
0 25 0 4.243 0.3038E+02 0.2130E+02 1.43 2
0 26 0 4.080 0.1054E+05 0.2527E+03 41.70 2
0 28 0 3.788 0.2113E+04 0.6034E+02 35.02 2
0 29 0 3.658 0.6292E+01 0.2727E+02 0.23 2
0 30 0 3.536 0.7334E+04 0.2572E+03 28.52 1
0 31 0 3.422 -0.4189E+02 0.5662E+02 -0.74 1
0 32 0 3.315 0.7962E+01 0.4740E+02 0.17 1
0 33 0 3.214 -0.1009E+02 0.5204E+02 -0.19 1
0 34 0 3.120 0.1827E+04 0.8498E+02 21.50 1
0 35 0 3.031 0.1701E+02 0.4512E+02 0.38 1
0 36 0 2.946 0.2340E+03 0.5634E+02 4.15 1
0 37 0 2.867 0.5759E+02 0.5532E+02 1.04 1
0 38 0 2.791 0.3205E+04 0.1241E+03 25.83 1
0 39 0 2.720 0.7106E+02 0.5549E+02 1.28 1
0 40 0 2.652 0.1882E+02 0.6010E+02 0.31 1
0 41 0 2.587 -0.4538E+02 0.5319E+02 -0.85 1
0 42 0 2.526 0.4779E+04 0.1783E+03 26.81 1
0 43 0 2.467 -0.1161E+02 0.6746E+02 -0.17 1
0 44 0 2.411 0.3388E+04 0.1429E+03 23.70 1
0 45 0 2.357 0.7914E+01 0.6885E+02 0.11 1
0 46 0 2.306 0.6401E+03 0.8085E+02 7.92 1
0 47 0 2.257 0.8448E+01 0.7337E+02 0.12 1
0 48 0 2.210 0.1121E+03 0.7723E+02 1.45 1
0 49 0 2.165 0.9636E+02 0.8220E+02 1.17 1
0 50 0 2.121 0.9795E+03 0.9043E+02 10.83 1
0 51 0 2.080 0.9658E+02 0.8250E+02 1.17 1
0 52 0 2.040 -0.1239E+02 0.8772E+02 -0.14 1
0 53 0 2.001 -0.4405E+01 0.8213E+02 -0.05 1
0 54 0 1.964 0.5261E+03 0.8460E+02 6.22 1
0 55 0 1.929 0.1024E+03 0.8495E+02 1.21 1
0 56 0 1.894 0.2097E+04 0.1220E+03 17.18 1
0 57 0 1.861 -0.9680E+02 0.8166E+02 -1.19 1
0 58 0 1.829 -0.1196E+03 0.8407E+02 -1.42 1
0 59 0 1.798 0.1302E+03 0.8407E+02 1.55 1
0 60 0 1.768 0.5786E+03 0.8353E+02 6.93 1
0 61 0 1.739 0.1614E+02 0.7909E+02 0.20 1
3 0 0 18.729 0.4087E+01 0.4877E+01 0.84 2
4 0 0 14.047 0.4620E+04 0.1563E+03 29.57 1
5 0 0 11.238 0.2628E+02 0.7980E+01 3.29 2
7 0 0 8.027 0.1388E+03 0.1199E+02 11.57 2
8 0 0 7.023 0.4997E+04 0.1207E+03 41.41 2
9 0 0 6.243 -0.3491E+01 0.1404E+02 -0.25 2
10 0 0 5.619 0.7205E+03 0.2461E+02 29.28 2
16 0 0 3.512 0.7500E+03 0.3599E+02 20.84 2
17 0 0 3.305 0.2017E+03 0.2954E+02 6.83 2
18 0 0 3.122 0.8955E+04 0.2177E+03 41.14 2
19 0 0 2.957 0.2619E+02 0.3592E+02 0.73 1
20 0 0 2.809 0.5455E+03 0.4308E+02 12.66 1
21 0 0 2.676 0.4300E+01 0.3632E+02 0.12 1
22 0 0 2.554 0.6695E+03 0.4925E+02 13.59 1
23 0 0 2.443 0.1916E+02 0.4766E+02 0.40 1
24 0 0 2.341 0.7016E+03 0.6038E+02 11.62 1
25 0 0 2.248 -0.6989E+02 0.5147E+02 -1.36 1
26 0 0 2.161 0.9047E+03 0.6482E+02 13.96 1
27 0 0 2.081 0.6900E+02 0.5447E+02 1.27 1
28 0 0 2.007 0.4734E+03 0.5668E+02 8.35 1
29 0 0 1.938 0.1150E+03 0.5947E+02 1.93 1
Res OBS UNIQUE POS COMP R(obs) R(exp) I/Sig Rmeas CC1/2
4.96 13512 3348 3568 93.8% 2.5% 2.5% 13461 47.52 2.8% 99.9* 2
0.859 2641
3.56 24267 5885 5938 99.1% 2.4% 2.5% 24249 48.26 2.7% 99.9*
-2 0.809 4704
2.93 22330 7401 7551 98.0% 2.8% 2.7% 22018 34.01 3.4% 99.8*
3 0.873 2097
2.54 19890 8751 8971 97.5% 3.5% 3.5% 19338 22.53 4.7% 99.6*
2 0.968 232
2.28 20645 9773 10080 97.0% 4.6% 4.7% 19775 16.89 6.2% 99.4*
-99 0.919 2
2.08 21352 10332 11093 93.1% 6.1% 6.5% 20047 12.69 8.3% 99.0*
0 0.000 0
1.93 21331 10559 12116 87.1% 9.4% 10.2% 19606 8.51 12.7% 97.7*
0 0.000 0
1.80 19417 9965 12977 76.8% 15.5% 17.2% 17229 5.19 21.1% 94.5*
0 0.000 0
1.70 11478 6792 13746 49.4% 22.2% 24.3% 8608 3.24 30.2% 88.2*
0 0.000 0
total174222 72806 86040 84.6% 3.5% 3.7% 164331 18.57 4.4% 99.8*
1 0.840 9676
Systematic absences suggest one 2(1) fold rotation symetry axis
Almost 2 more but there are a number of violations.
Very low res data complete to ~82% (10-20A)
twinning analysis
Accentric
<I**2>/<I>**2 = 1.97 (untwinned 2.0, pefect twin 1.5)
<F>**2/<F**2> =0.792 (0.785 untwinned, 0.885 perfect twin)
<|L|> = 0.490 (0.5 expected, 0.375, perfect twin)
<L**2> = 0.324 (0.33 expected, perfect twin .20)
possible twin law h, -k, -l
Wilson plot - estimated B factor = 10.6
2 molec /asu
1 off origin patterson peak Height: 4.974
Molecular Replacement (Phaser, automated) and refinement
Solution in P21 when processed in P2
Refinement without twin law:
Rwork = 0.3387
Rfree = 0.3650
~85% Ramachandran favored (+ missing part of helix)
Refinement with twin law:
Rwork = 0.2994
Rfree = 0.3214
--
Yarrow Madrona
Postdoctoral Scholar
Ortiz De Montellano Lab
Pharmeceutical Chemistry Dept.
University of California, San Francisco
Genentech Hall, Rm N556
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