Yes - convincing.. Follow Jacob's advice and see how the maps look. Eleanor
On 15 April 2017 at 06:04, Alex Lee <alexlee198...@gmail.com> wrote: > Thanks Eleanor, > If I understand right, there is just 1 TFZ. TFZ== means "Translation > Function Z-score equivalent, only calculated for the top solution after > refinement (or for the number of top files specified by TOPFILES)" so there > could be many TFZ==. > > Twinning analysis attached below: > > TWINNING ANALYSIS: > > > Global twinning statistics. > > > These tests rely on the fact that it is highly improbably that very weak or > very strong reflections will coincide, therefore, the tails for the > distribution of twinned datasets will be less pronounced > > > Data truncated to 67.93 - 3.50 A resolution > > $TABLE: Cumulative intensity distribution: > > $GRAPHS: Cumulative intensity distribution (Acentric and > centric):N:1,2,3,4,5,6: > > $$ Z Acent_theor Acent_twin Acent_obser Cent_theor Cent_obser $$ > > $$ > > 0.00000 0.00000 0.00000 0.01988 0.00000 - > > 0.04000 0.03921 0.00303 0.03225 0.15852 - > > 0.08000 0.07688 0.01151 0.04745 0.22270 - > > 0.12000 0.11308 0.02458 0.06655 0.27097 - > > 0.16000 0.14786 0.04148 0.09008 0.31084 - > > 0.20000 0.18127 0.06155 0.11562 0.34528 - > > 0.24000 0.21337 0.08420 0.14170 0.37579 - > > 0.28000 0.24422 0.10891 0.16750 0.40330 - > > 0.32000 0.27385 0.13524 0.19450 0.42839 - > > 0.36000 0.30232 0.16279 0.22295 0.45149 - > > 0.40000 0.32968 0.19121 0.25213 0.47291 - > > 0.44000 0.35596 0.22021 0.28103 0.49288 - > > 0.48000 0.38122 0.24953 0.30678 0.51158 - > > 0.52000 0.40548 0.27895 0.33697 0.52916 - > > 0.56000 0.42879 0.30829 0.36569 0.54574 - > > 0.60000 0.45119 0.33737 0.39332 0.56142 - > > 0.64000 0.47271 0.36607 0.41967 0.57629 - > > 0.68000 0.49338 0.39428 0.44513 0.59041 - > > 0.72000 0.51325 0.42190 0.47060 0.60386 - > > 0.76000 0.53233 0.44885 0.49430 0.61667 - > > 0.80000 0.55067 0.47507 0.51550 0.62891 - > > 0.84000 0.56829 0.50052 0.53736 0.64060 - > > 0.88000 0.58522 0.52516 0.55780 0.65180 - > > 0.92000 0.60148 0.54896 0.57822 0.66253 - > > 0.96000 0.61711 0.57191 0.59779 0.67281 - > > 1.00000 0.63212 0.59399 0.61638 0.68269 - > > 1.04000 0.64655 0.61521 0.63496 0.69218 - > > 1.08000 0.66040 0.63557 0.65120 0.70130 - > > 1.12000 0.67372 0.65507 0.66731 0.71008 - > > 1.16000 0.68651 0.67373 0.68115 0.71853 - > > 1.20000 0.69881 0.69156 0.69557 0.72668 - > > 1.24000 0.71062 0.70857 0.70990 0.73453 - > > 1.28000 0.72196 0.72480 0.72386 0.74210 - > > 1.32000 0.73286 0.74025 0.73801 0.74941 - > > 1.36000 0.74334 0.75495 0.74895 0.75646 - > > 1.40000 0.75340 0.76892 0.75913 0.76328 - > > 1.44000 0.76307 0.78220 0.77024 0.76986 - > > 1.48000 0.77236 0.79480 0.78058 0.77623 - > > 1.52000 0.78129 0.80675 0.79061 0.78238 - > > 1.56000 0.78986 0.81807 0.79977 0.78833 - > > 1.60000 0.79810 0.82880 0.80902 0.79410 - > > 1.64000 0.80602 0.83895 0.81795 0.79967 - > > 1.68000 0.81363 0.84855 0.82691 0.80508 - > > 1.72000 0.82093 0.85763 0.83511 0.81031 - > > 1.76000 0.82796 0.86621 0.84245 0.81538 - > > 1.80000 0.83470 0.87431 0.85141 0.82029 - > > 1.84000 0.84118 0.88196 0.85966 0.82505 - > > 1.88000 0.84741 0.88917 0.86721 0.82967 - > > 1.92000 0.85339 0.89597 0.87415 0.83414 - > > 1.96000 0.85914 0.90238 0.87977 0.83849 - > > 2.00000 0.86466 0.90842 0.88629 0.84270 - > > $$ > > > > The culmulative intensity, N(Z), plot is diagnostic for both twinning and > tNCS. For twinned data there are fewer weak reflections, therefore, N(Z) is > sigmoidal for twinned data. However, if both twinning and tNCS are present, > the effects may cancel each out. Therefore the results of the L-test and > patterson test should be consulted > > > > L test for twinning: (Padilla and Yeates Acta Cryst. D59 1124 (2003)) > > L statistic = 0.416 (untwinned 0.5 perfect twin 0.375) > > Data has used to 67.93 - 3.50 A resolution > > Relation between L statistics and twinning fraction: > > Twinning fraction = 0.000 L statistics = 0.500: > > Twinning fraction = 0.100 L statistics = 0.440: > > Twinning fraction = 0.500 L statistics = 0.375: > > > The L test suggests data is twinned > > All data regardless of I/sigma(I) has been included in the L test > > > > $TABLE: L test for twinning: > > $GRAPHS: cumulative distribution function for |L|, twin fraction of > 0.18:0|1x0|1:1,2,3,4: > > $$ |L| N(L) Untwinned Twinned $$ > > $$ > > 0.0000 0.0000 0.0000 0.0000 > > 0.0500 0.0666 0.0500 0.0749 > > 0.1000 0.1339 0.1000 0.1495 > > 0.1500 0.2007 0.1500 0.2233 > > 0.2000 0.2648 0.2000 0.2960 > > 0.2500 0.3263 0.2500 0.3672 > > 0.3000 0.3870 0.3000 0.4365 > > 0.3500 0.4475 0.3500 0.5036 > > 0.4000 0.5070 0.4000 0.5680 > > 0.4500 0.5646 0.4500 0.6294 > > 0.5000 0.6206 0.5000 0.6875 > > 0.5500 0.6756 0.5500 0.7418 > > 0.6000 0.7274 0.6000 0.7920 > > 0.6500 0.7768 0.6500 0.8377 > > 0.7000 0.8229 0.7000 0.8785 > > 0.7500 0.8643 0.7500 0.9141 > > 0.8000 0.9025 0.8000 0.9440 > > 0.8500 0.9355 0.8500 0.9679 > > 0.9000 0.9628 0.9000 0.9855 > > 0.9500 0.9842 0.9500 0.9963 > > 1.0000 1.0000 1.0000 1.0000 > > $$ > > > > The Cumulative |L| plot for acentric data, where L = (I1-I2)/(I1+I2). This > depends on the local difference in intensities. The difference operators > used link to the neighbouring reflections taking into account possible tNCS > operators. > > Note that this estimate is not as reliable as obtained via the H-test or ML > Britton test if twin laws are available. However, it is less prone to the > effects of anisotropy than the H-test > > > Reference: Padilla, Yeates. A statistic for local intensity differences: > robustness to anisotropy and pseudo-centering and utility for detecting > twinning. Acta Cryst. D59, 1124-30, 2003. > > > > Mean acentric moments I from input data: > > > <I^2>/<I>^2 = 1.818 (Expected = 2.000, Perfect Twin = 1.500) > > <I^3>/<I>^3 = 5.843 (Expected value = 6.000, Perfect Twin = 3.000) > > <I^4>/<I>^4 = 37.521 (Expected value = 24.000, Perfect Twin = 7.500) > > > $TABLE: Acentric Moments of I: > > $GRAPHS: 2nd moment of I 1.818 (Expected value = 2, Perfect Twin = > 1.5):0|0.112x0|5:1,2: > > : 3rd & 4th Moments of I (Expected values = 6, 24, Perfect twin = 3, > 7.5):0|0.112x0|36:1,3,4: > > $$ 1/resol^2 <I**2> <I**3> <I**4> $$ > > $$ > > 0.006970 3.366 30.978 430.240 > > 0.015047 1.914 5.340 18.196 > > 0.021148 1.777 4.490 14.406 > > 0.026505 1.960 6.778 34.617 > > 0.031327 1.705 4.206 14.007 > > 0.035779 1.695 3.932 11.011 > > 0.039978 1.912 5.494 19.767 > > 0.043990 1.720 3.990 11.197 > > 0.047857 1.787 5.324 23.536 > > 0.051571 1.892 5.798 24.313 > > 0.055079 1.668 4.328 16.686 > > 0.058588 1.936 6.063 25.829 > > 0.061936 1.595 3.392 8.645 > > 0.065227 1.578 3.338 8.668 > > 0.068297 1.628 3.610 10.245 > > 0.071442 1.633 3.616 9.824 > > 0.074425 1.502 2.992 7.248 > > 0.077502 1.607 3.465 9.355 > > 0.080350 1.609 3.327 7.939 > > 0.083247 1.634 3.766 11.561 > > 0.086126 1.845 5.379 21.920 > > 0.088817 1.455 2.697 5.933 > > 0.091594 2.329 13.127 120.470 > > 0.094336 2.184 7.879 36.506 > > 0.096883 2.403 8.850 39.439 > > 0.099563 1.963 5.747 21.618 > > 0.102142 1.753 4.213 12.919 > > 0.104702 1.762 3.872 10.083 > > 0.107158 1.816 4.074 10.915 > > 0.109761 1.657 3.348 7.942 > > 0.112129 1.864 4.539 13.753 > > $$ > > > First principles calculation has found 3 potential twinning operators > > > # twinning operator score type > > 0 k,h,-l 0.00 pm > > 1 -h,-k,l 0.00 pm > > 2 -h,h+k,-l 0.00 pm > > m merohedral > > pm pseudo-merohedral > > The score gives an indication of the closure of the twinning operation. The > lower the values > > the more higher the overlap. > > The appearance of twinning operators only indicates that the crystal symmetry > and lattice symmetry permit twinning. It does not mean that there is > twinning present. Only the presence of statistics consistent with twinning > gives a strong indicator. > > > Twinning operator based tests: > > > H-test: Cumulative plot of H=|I-T(I)|/(I-T(I)) for twin related reflections. > This should be linear with slope 1/(1-2a). > > > > $TABLE: H test for twinning > > $GRAPHS: cumulative distribution function for |H| (operator k, h, -l) alpha = > 0.42:0|1x0|1:1,2,3,4,5,6,7: > > : cumulative distribution function for |H| (operator -h, -k, l) alpha = > 0.39:0|1x0|1:1,2,3,4,5,6,8: > > : cumulative distribution function for |H| (operator -h, h+k, -l) alpha = > 0.39:0|1x0|1:1,2,3,4,5,6,9: > > $$ |H| 0.4 0.3 0.2 0.1 0.0 k,h,-l -h,-k,l -h,h+k,-l$$ > > $$ > > 0.00 0.0 0.0 0.0 0.0 0.0 0.00 0.00 0.00 > > 0.05 - - - - - 0.68 0.43 0.45 > > 0.10 - - - - - 0.88 0.72 0.72 > > 0.15 - - - - - 0.94 0.86 0.86 > > 0.20 - - - - - 0.96 0.92 0.93 > > 0.25 - - - - - 0.98 0.95 0.96 > > 0.30 - - - - - 0.98 0.97 0.97 > > 0.35 - - - - - 0.99 0.98 0.98 > > 0.40 - - - - - 0.99 0.98 0.98 > > 0.45 - - - - - 0.99 0.99 0.99 > > 0.50 - - - - - 1.00 0.99 0.99 > > 0.55 - - - - - 1.00 0.99 0.99 > > 0.60 - - - - - 1.00 0.99 0.99 > > 0.65 - - - - - 1.00 1.00 1.00 > > 0.70 - - - - - 1.00 1.00 1.00 > > 0.75 - - - - - 1.00 1.00 1.00 > > 0.80 - - - - - 1.00 1.00 1.00 > > 0.85 - - - - - 1.00 1.00 1.00 > > 0.90 - - - - - 1.00 1.00 1.00 > > 0.95 - - - - - 1.00 1.00 1.00 > > 1.00 5.0 2.5 1.67 1.25 1.0 1.00 1.00 1.00 > > $$ > > > Britton plot: Plot of number of negative detwinned intensities. > > > > $TABLE: Britton plot for twinning > > $GRAPHS: aI1+(1-a)I2 > 0 (operator k, h, -l) alpha = 0.39:A:1,2: > > : aI1+(1-a)I2 > 0 (operator -h, -k, l) alpha = 0.37:A:1,3: > > : aI1+(1-a)I2 > 0 (operator -h, h+k, -l) alpha = 0.37:A:1,4: > > $$ alpha k,h,-l -h,-k,l -h,h+k,-l$$ > > $$ > > 0.00 0.00 0.00 0.00 > > 0.03 0.00 0.00 0.00 > > 0.05 0.00 0.00 0.00 > > 0.07 0.00 0.00 0.00 > > 0.10 0.00 0.00 0.00 > > 0.12 0.00 0.00 0.00 > > 0.15 0.00 <B><FONT COLOR='#FF0000'> 0.00 0.00 > > 0.17 0.00 0.00 0.00 > > 0.20 0.00 0.00 0.00 > > 0.23 0.00 0.00 0.00 > > 0.25 0.00 0.00 0.00 > > 0.28 0.00 0.00 0.00 > > 0.30 0.00 0.01 0.01 > > 0.33 0.00 0.01 0.01 > > 0.35 0.01 0.01 0.01 > > 0.38 0.01 0.01 0.01 > > 0.40 0.01 0.02 0.02 > > 0.42 0.02 0.04 0.04 > > 0.45 0.03 0.07 0.07 > > 0.47 0.08 0.14 0.14 > > $$ > > > ML-Britton: Plot of number of negative detwinned intensities. The ML element > corrects for the sigma in the observed intensity and for the effects of a > single tNCS operator, if it is present. > > > > $TABLE: ML-Britton test for twinning > > $GRAPHS: aI1+(1-a)I2 > 0 (operator k, h, -l) alpha = 0.47:A:1,2: > > : aI1+(1-a)I2 > 0 (operator -h, -k, l) alpha = 0.45:A:1,3: > > : aI1+(1-a)I2 > 0 (operator -h, h+k, -l) alpha = 0.45:A:1,4: > > $$ alpha k,h,-l -h,-k,l -h,h+k,-l$$ > > $$ > > 0.00 0.26 0.26 0.26 > > 0.03 -1922.56 -1919.46 -1918.94 > > 0.05 -3987.00 -3977.78 -3974.50 > > 0.07 -6157.25 -6136.57 -6131.95 > > 0.10 -8447.11 -8411.39 -8406.03 > > 0.12 -10877.36 -10824.58 -10818.17 > > 0.15 -13468.83 -13394.48 -13385.80 > > 0.17 -16243.41 -16140.40 -16129.30 > > 0.20 -19226.63 -19084.18 -19071.17 > > 0.23 -22450.29 -22252.97 -22240.06 > > 0.25 -25953.54 -25681.19 -25672.06 > > 0.28 -29785.11 -29408.61 -29408.35 > > 0.30 -34009.00 -33478.99 -33493.97 > > 0.33 -38707.08 -37931.58 -37972.35 > > 0.35 -43982.57 -42782.44 -42863.82 > > 0.38 -49953.87 -47966.38 -48105.22 > > 0.40 -56711.13 -53196.56 -53411.44 > > 0.42 -64141.85 -57712.76 -58018.04 > > 0.45 -71453.32 -60141.97 -60541.82 > > 0.47 -76835.02 -59398.04 -59888.16 > > $$ > > > Twin fraction estimates based on global statistics: > > Twin fraction estimate from L-test: 0.18 > > Twin fraction estimate from moments: 0.10 > > > Twin fraction estimates by twinning operator > > > The following operator based twinning estimates analyse data with each of the > possible twin operators. If twinning is present the most likely operator > will have a low RTwin score (<I-T(I)>/<I+T(I)>) and estimates of the twin > fraction above 0. > > > ------------------------------------------------------------------------------------------------- > > | operator | L-test | |Rtwin| | H-test | > Britton | ML Britton | > > ------------------------------------------------------------------------------------------------- > > | k, h, -l | Yes | 0.05 | 0.42 | > 0.39 | 0.47 ( N/A ) | > > | -h, -k, l | Yes | 0.08 | 0.39 | > 0.37 | 0.45 ( N/A ) | > > | -h, h+k, -l | Yes | 0.08 | 0.39 | > 0.37 | 0.45 ( N/A ) | > > ------------------------------------------------------------------------------------------------- > > > TWINNING SUMMARY > > > Twinning fraction from H-test: 0.42 > > Twinning fraction from L-Test: 0.18 > > > It is highly probable that your crystal is TWINNED. > > > Please use twin refinement after your model is almost completed and R-free > is below 40% > > > > On Fri, Apr 14, 2017 at 12:10 PM, Eleanor Dodson < > eleanor.dod...@york.ac.uk> wrote: > >> That twin factor list means the apparent crystal symmetry must be P6/mmm. >> >> You say you only have 2 molecules in the asymmetric unit of P32,therefor >> there must only be one in SGs P32 21 P32 12 >> >> So I dont understand why you have PHASER results like this: >> >> SOLU SET RFZ=4.4 TFZ=7.7 PAK=0 LLG=55 TFZ==9.6 LLG=350 TFZ==20.5 PAK=0 >> LLG=350 TFZ==20.5.. >> >> >> Why so many TFZ here - is that achieved after refinement or something? >> >> >> Eleanor >> >> >> And what does the twinning analysis suggest? >> >> >> >> >> On 14 April 2017 at 17:42, Keller, Jacob <kell...@janelia.hhmi.org> >> wrote: >> >>> As I mentioned off-list, it would be helpful to know how many types of >>> search models you are searching with—how many different molecules are in >>> the complex? It’s hard to interpret MR results otherwise. >>> >>> >>> >>> Also, since the higher-symmetry SG works in MR, you should try to refine >>> the model in that SG, with only two twin domains, refining twin fraction. I >>> can guarantee that a good reviewer will have you do this (if not, then not >>> a “good reviewer.”) >>> >>> >>> >>> JPK >>> >>> >>> >>> *From:* CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] *On Behalf >>> Of *Alex Lee >>> *Sent:* Friday, April 14, 2017 11:50 AM >>> >>> *To:* CCP4BB@JISCMAIL.AC.UK >>> *Subject:* Re: [ccp4bb] Refmac5 twin refinement pushing Rfree >>> surprisingly down >>> >>> >>> >>> Thanks Eleanor, I tried MR for P32 21 and P32 12. >>> >>> SG P3221: SOLU SET RFZ=5.3 TFZ=8.8 PAK=0 LLG=121 TFZ==11.2 LLG=944 >>> TFZ==29.2 PAK=0 LLG=944 TFZ==29.2 >>> >>> SOLU SPAC P 32 2 1 >>> >>> >>> >>> SG P3212: >>> >>> Solution #1 annotation (history): >>> >>> >>> >>> SOLU SET RFZ=4.4 TFZ=7.7 PAK=0 LLG=55 TFZ==9.6 LLG=350 TFZ==20.5 PAK=0 >>> LLG=350 TFZ==20.5 >>> >>> >>> >>> SOLU SPAC P 32 1 2 >>> >>> >>> >>> SG P32 >>> >>> SOLU SET RFZ=7.4 TFZ=10.4 PAK=0 LLG=187 TFZ==10.7 RF++ TFZ=17.0 PAK=0 >>> LLG=436 TFZ==17.8 LLG=1715 TFZ==34.3 PAK=0 >>> >>> LLG=1715 TFZ==34.3 >>> >>> SOLU SPAC P 32 >>> >>> >>> >>> Based on TFZ and LLG, the P32 seems to be best. But I'll also try to refine >>> and build P32 2 1 latter >>> >>> >>> >>> On Fri, Apr 14, 2017 at 4:32 AM, Eleanor Dodson < >>> eleanor.dod...@york.ac.uk> wrote: >>> >>> First - four way twinning is possible but pretty rare for macromolecules >>> >>> >>> >>> Pointless gives a very useful table of the CC agreement for each >>> possible symmetry operator individually. >>> >>> In this case with only two molecules in the asymmetric unit you you >>> could only have a higher symmetry SG as >>> >>> P32 21 P32 12 or P64 >>> >>> >>> >>> These would require as symmetry operators - >>> >>> P32 21 - a three fold and a two fold k h -l >>> >>> P32 12 - a three fold and a two fold -k -h -l >>> >>> >>> >>> P64 - a six fold >>> >>> >>> >>> If the scores for one set are better than the others you probably have >>> that SG >>> >>> >>> >>> However high degrees of twinning can disguise the symmetry scores of >>> course.. >>> >>> >>> >>> >>> >>> >>> >>> On 14 April 2017 at 04:46, Keller, Jacob <kell...@janelia.hhmi.org> >>> wrote: >>> >>> Try MR with one copy in all space groups of PG 321/312 using Phaser. >>> Going from PG 3 to PG 32 should halve the number of copies per ASU. You may >>> have to re-process your data in the higher point group to do this. >>> >>> >>> >>> Or you might actually have a tetartohedral twin, but just try with the >>> higher-symmetry point group first, see what happens. >>> >>> >>> >>> JPK >>> >>> >>> >>> *From:* Alex Lee [mailto:alexlee198...@gmail.com] >>> *Sent:* Thursday, April 13, 2017 11:32 PM >>> >>> >>> *To:* Keller, Jacob <kell...@janelia.hhmi.org> >>> *Cc:* CCP4BB@JISCMAIL.AC.UK >>> *Subject:* Re: [ccp4bb] Refmac5 twin refinement pushing Rfree >>> surprisingly down >>> >>> >>> >>> Hi Keller, >>> >>> >>> >>> Thanks for the suggestions! I only have two copies in ASU at SG P32. >>> Zanuda also suggests P32 is the best SG. >>> >>> >>> >>> On Thu, Apr 13, 2017 at 8:12 PM, Keller, Jacob <kell...@janelia.hhmi.org> >>> wrote: >>> >>> Yes, this was my case exactly—it looks like there are two pairs of >>> coupled twin domains: a,c and b,d. Assuming you have multiple copies of >>> your model in the same ASU, try doing MR in higher symmetry space groups of >>> point group 312 or 321, like P3212 etc. There is this handy page with all >>> the space groups and their possible twin operators: >>> http://www.ccp4.ac.uk/html/twinning.html. >>> >>> >>> >>> The twin fractions indicate a high twin fraction—~46% if actually >>> hemihedral! >>> >>> >>> >>> Also take a look at the paper I referenced for more info. I can send you >>> a .pdf if you need me to. >>> >>> >>> >>> Please let me know how it works out—I am interested in these types of >>> things! >>> >>> >>> >>> JPK >>> >>> >>> >>> *From:* Alex Lee [mailto:alexlee198...@gmail.com] >>> *Sent:* Thursday, April 13, 2017 9:08 PM >>> *To:* Keller, Jacob <kell...@janelia.hhmi.org> >>> *Cc:* CCP4BB@JISCMAIL.AC.UK >>> >>> >>> *Subject:* Re: [ccp4bb] Refmac5 twin refinement pushing Rfree >>> surprisingly down >>> >>> >>> >>> Hi Keller, >>> >>> >>> >>> I do not how to check twin fraction after Refmac (I guess it's somewhere >>> in log file). From the log file it seems I have four twin domain: >>> >>> Twin operators with estimated twin fractions **** >>> >>> >>> >>> Twin operator: H, K, L: Fraction = 0.275; Equivalent operators: K, >>> -H-K, L; -H-K, H, L >>> >>> Twin operator: -K, -H, -L: Fraction = 0.228; Equivalent operators: -H, >>> H+K, -L; H+K, -K, -L >>> >>> Twin operator: K, H, -L: Fraction = 0.270; Equivalent operators: H, >>> -H-K, -L; -H-K, K, -L >>> >>> Twin operator: -H, -K, L: Fraction = 0.228; Equivalent operators: -K, >>> H+K, L; H+K, -H, L >>> >>> >>> >>> On Thu, Apr 13, 2017 at 4:36 PM, Keller, Jacob <kell...@janelia.hhmi.org> >>> wrote: >>> >>> What was the refined twin fraction after Refmac? It’s much more accurate >>> than initial tests. Also, how many twin domains do you have? If you have >>> many, it might be a higher space group but with less twinning. I recently >>> had a case in which apparent tetartohedral (four-domain) twinning in P32 >>> was really hemihedral (two-domain) twinning in P3212: >>> >>> >>> >>> *Acta Cryst. <http://journals.iucr.org/d>* (2017). D*73* >>> <http://journals.iucr.org/d/contents/backissues.html>, 22-31 >>> https://doi.org/10.1107/S2059798316019318 >>> >>> >>> >>> Jacob >>> >>> >>> >>> *From:* CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] *On Behalf >>> Of *Eleanor Dodson >>> *Sent:* Thursday, April 13, 2017 3:11 PM >>> *To:* CCP4BB@JISCMAIL.AC.UK >>> *Subject:* Re: [ccp4bb] Refmac5 twin refinement pushing Rfree >>> surprisingly down >>> >>> >>> >>> Twin refinement cannot be compared directly to untwinned - the R factors >>> are between different parameters - without twinning it is assumed you have >>> an amplitude obtained more or less from sqrt(I But for a twinned data set >>> that I is actually [ I1 + twin_factor I2 ] so the amplitude is not really >>> correct and twinned refinement will give a much better estimate. >>> >>> >>> >>> However you need to be careful that you have assigned the same FreeR >>> flag to reflection pair related by the twin law. The modern program in the >>> CCP4 data reduction pipeline looks after this pretty automatically - all >>> possible symmetry equivalents are assigned the same FreeR but older >>> software did not do this.. >>> >>> >>> >>> You can check it by looking at some twin equivalents - in SG P32 these >>> could be h k l and -h, -k, l or h k l and k h -l or h k l and -k, -h, -l . >>> >>> >>> >>> Ideally they all should have the same Free R flag.. >>> >>> >>> >>> Eleanor >>> >>> >>> >>> PS - the acid test is: Do the maps look better? >>> >>> >>> >>> E >>> >>> >>> >>> >>> >>> On 13 April 2017 at 19:52, Robbie Joosten <r.joos...@nki.nl> wrote: >>> >>> Hi Alex, >>> >>> >>> >>> You are not giving the number after refinement without the twin >>> refinement. Nevertheless, R-free drops like this are not unheard of. You >>> should check your Refmac log file, it would warn you of potential space >>> group errors. Refmac will also give you a refined estimate of the twin >>> fraction. >>> >>> >>> >>> Cheers, >>> >>> Robbie >>> >>> >>> >>> Sent from my Windows 10 phone >>> >>> >>> >>> *Van: *Alex Lee <alexlee198...@gmail.com> >>> *Verzonden: *donderdag 13 april 2017 19:19 >>> *Aan: *CCP4BB@JISCMAIL.AC.UK >>> *Onderwerp: *[ccp4bb] Refmac5 twin refinement pushing Rfree >>> surprisingly down >>> >>> >>> >>> Dear All, >>> >>> >>> >>> I have a protein/dna complex crystal and data collected at 3A and >>> another set at 2.8A, space group P32. L test shows twinning (fraction >>> around 0.11). The structure solved by MR and model building of the complex >>> finish (no solvent built yet, I do not think it's good to build solvent in >>> such low resolution data). >>> >>> >>> >>> I did Refmac5 to refine my structure (restraint refinement) with or >>> without twinning, to my surprise, the Rfree drops a lot after twin >>> refinement of two data sets. Summary below: >>> >>> >>> >>> 2.8A dataset: before twin refine 34%, 29%; after twin refine:24%, 19% >>> >>> 3A dataset: before twin refine 30%;26%; after refine 25%, 18% >>> >>> >>> >>> I know that a lot of threads in CCP4bb talking about Rfree after twin >>> refine and Rfree without twin refine can not compare directly. By drop R >>> free this much by twin refine, it gives me a feeling of too good to be true >>> (at such low resolution with such good Rfree, maybe overrefined a lot?), >>> but from the density map after twin refine, it does seem better than no >>> twin refine map. >>> >>> >>> >>> I do not know if reviewers are going to challenge this part. >>> >>> >>> >>> Any input is appreciated. >>> >>> >>> >>> >>> >>> >>> >>> >>> >>> >>> >>> >>> >>> >>> >>> >>> >> >> >
