Hi, There
1.I have collected two data sets one for the native protein (60Kda) and complex of native with other protein of 8 Kda. The shape of native dataset is cubic while that of the complexed one is triangular. A number of dataset were collected for these two and i successfully indexed them to a spacegroup of C2.
2.While doing molecular replacement for the native data using a monomeric template clear solution with 3 monomers in the asyymetric unit was obtained and structure solved successfully with R/rfee of 19/23 at 2.5A reolution.
3.The molecular replacement of the complexed dataset with the same template gave a solution with 2 monomers in asymmetric unit with a lot of clashes between the two subnuints. when i closely examined the logfile i observed that there was pseudotranslation detected by the program in all of my complex dataset. I run xtiage as well as pointless and truncate which rules out any twinning as well as wrong space group assignment. Pseudotranslation of 18% has been detected by balbes.
4. I also run the Balbes at Murshodov server which gave a solution with 4 monomers in the asymmetric but still showing a lot of clashes in the asymmetric unit (R/Rfree 0.43/0.47) followed by automatic refinemnet with Arp/wARp which gave a model . When i examine the density of the model i saw a lot of clashes in the asymmetric unit but the density are interpretable so that bakbone can be easily modelled. But the R and R free donot goes down from (R/Rfree 0.43/0.47)
I have pasted the logfile below from molrep for your convenience. Kindly suggest me how can i proceed with this dataset. That will be very kind of you if you spend some time over my problem. Another question , is it possible that this pseudotranslation is due to the smaller protein of the complex which has slightly changed the crystal packing. ( SDS done on the complex crystals clearly showed the presence of both the proteins).
--- Check Patterson for pseudo-translation ---
PST_limit : 0.125 of origin peak
INFO: pseudo-translation was detected.
Origin Patterson peak: P,P/sig : 80563.180 266.237
1 Patterson. peak : p,P/sig : 80560.719 266.228
2 Patterson peak : P,P/sig : 13992.708 46.242
3 Patterson peak : P,P/sig : 13992.708 46.242
Peak 1: trans.vector /ort/ : 60.164 98.423 0.000
trans.vector /frac/: 0.500 0.500 0.000
Peak 2: trans.vector /ort/ : 31.861 0.000 0.000
trans.vector /frac/: 0.265 0.000 0.000
Peak 3: trans.vector /ort/ : 28.303 98.423 0.000
trans.vector /frac/: 0.235 0.500 0.000
INFO: translation vector of peak 2 will be used.
WARNING: with keyword: PST --> MODE = F , STICK = N
Sol_ Space group : C 1 2 1
Sol_ No: 5 Sett: 2
Sol_ Cell: 120.328 196.846 109.285 90.00 113.84 90.00
Sol_--- Rotation function ---
Sol_
Radius of gyration : 23.58
Sol_ Radius of integration : 47.16
Sol_ Resmin,Resmax : 48.03 2.42
WARNING: For this radius integration program uses data included
between 48.0 and 2.71 (angstrom..)
--- rfcoef for model ---
--- rfcoef for Fobs ---
NCS (from Self rotation Function): 1
NCS_model (from Model Self rotation Function): 1
Program will use NCS_model =: 1
Number of RF peaks : 30
theta phi chi alpha beta gamma Rf Rf/sigma
Sol_RF 1 106.45 50.40 1.37 320.21 1.31 39.41 0.9325E+05 19.98
Sol_RF 2 0.00 0.00 0.00 0.00 0.00 0.00 0.8668E+05 18.57
Sol_RF 3 158.10 -1.79 120.96 209.61 37.87 33.18 0.8147E+05 17.46
Sol_RF 4 22.21 -179.07 120.40 149.19 38.30 327.33 0.7596E+05 16.28
Sol_RF 5 0.83 144.00 179.87 143.94 1.66 35.93 0.4234E+05 9.07
Sol_RF 6 146.73 90.95 73.37 329.03 38.27 327.14 0.3574E+05 7.66
Sol_RF 7 0.00 0.00 179.92 97.16 0.00 82.76 0.3504E+05 7.51
Sol_RF 8 34.01 88.19 71.45 29.00 38.12 32.61 0.3354E+05 7.19
Sol_RF 9 77.12 174.71 90.84 97.45 87.95 288.04 0.2136E+05 4.58
Sol_RF 10 134.42 132.29 7.11 39.81 5.07 315.22 0.2089E+05 4.48
Sol_RF 11 152.93 177.57 136.00 21.98 49.92 206.84 0.1861E+05 3.99
Sol_RF 12 156.31 -169.21 19.45 91.88 7.78 250.29 0.1849E+05 3.96
Sol_RF 13 130.28 116.07 92.63 351.98 66.96 299.83 0.1810E+05 3.88
Sol_RF 14 0.00 0.00 11.30 11.30 0.00 0.00 0.1808E+05 3.87
Sol_RF 15 36.79 -89.11 90.53 219.84 50.36 218.06 0.1801E+05 3.86
Sol_RF 16 134.90 -102.85 173.11 82.03 90.00 107.72 0.1761E+05 3.77
Sol_RF 17 106.06 0.94 54.00 262.92 51.73 81.03 0.1753E+05 3.76
Sol_RF 18 135.11 85.04 162.00 277.64 88.39 287.56 0.1728E+05 3.70
Sol_RF 19 127.04 109.26 97.92 344.58 74.03 306.06 0.1722E+05 3.69
Sol_RF 20 141.26 -144.07 13.23 120.76 8.27 228.90 0.1705E+05 3.65
Sol_RF 21 142.16 -164.54 114.88 54.42 62.26 203.50 0.1675E+05 3.59
Sol_RF 22 137.57 -150.27 12.10 115.26 8.16 235.79 0.1657E+05 3.55
Sol_RF 23 162.01 178.37 129.40 24.79 32.42 208.06 0.1654E+05 3.54
Sol_RF 24 135.22 28.82 14.02 293.83 9.86 56.20 0.1636E+05 3.51
Sol_RF 25 134.21 84.96 161.20 278.32 90.00 288.40 0.1621E+05 3.47
Sol_RF 26 43.85 57.57 90.92 3.81 59.17 68.66 0.1616E+05 3.46
Sol_RF 27 22.55 173.85 110.70 137.04 36.79 329.34 0.1615E+05 3.46
Sol_RF 28 142.62 94.57 75.18 333.11 43.47 323.97 0.1612E+05 3.46
Sol_RF 29 25.92 179.07 113.06 142.75 42.77 324.61 0.1609E+05 3.45
Sol_RF 30 128.76 -103.39 65.70 144.60 50.05 171.38 0.1599E+05 3.43
Final number of peaks : 30
INFO: Relations between peaks see in molrep.doc
Time: 22h 12m 3s Elapsed: 0h 3m 46s
Sol_
Sol_--- Translation function ---
Sol_ Resmin,Resmax : 48.03 2.42
Sol_ NCS model : 1
--- List of psewdo-translation vectors ---
PST_vector 1: 0.265 0.000 0.000
---
Time_elapsed: 0h 4m 1s Remained: 0h 7m 15s
Sol_ RF TF Tf/sig TFcntrst PFind PF PFmin wRfac Scor Scor_max Cntrst
Sol___1__1 24.83 10.203 1.00 1.00 -4.18 0.595 0.390 0.390 0.00
Sol___2__1 21.58 11.436 1.00 1.00 -4.13 0.601 0.377 0.390 0.00
Sol___3_14 4.093 4.995 1.00 1.00 -3.57 0.614 0.353 0.390 0.00
Sol___4__1 21.04 13.251 1.00 0.98 -3.46 0.607 0.361 0.390 0.00
Sol___5__1 14.68 3.760 1.00 1.00 -3.25 0.614 0.349 0.390 3.99
Sol___6__6 5.017 0.269 1.00 1.00 -4.19 0.625 0.325 0.390 3.14
Sol___7__1 13.18 3.772 1.00 1.00 -3.18 0.617 0.342 0.390 3.14
Sol___8__6 4.372 2.119 1.00 1.00 -3.86 0.622 0.332 0.390 3.02
Sol___9__3 4.122 2.055 1.00 1.00 -3.41 0.626 0.318 0.390 3.20
Sol__10__2 4.511 1.500 1.00 1.00 -3.91 0.627 0.317 0.390 2.96
Sol__11__2 4.970 2.936 1.00 1.00 -3.99 0.628 0.316 0.390 2.79
Sol__12__5 3.856 1.531 1.00 1.00 -3.73 0.628 0.313 0.390 3.01
Sol__13__6 3.437 1.204 1.00 1.00 -4.07 0.629 0.315 0.390 2.92
INFO: contrast is good enough. Stop this run
--- Summary ---
S_ RF TF theta phi chi tx ty tz TFcnt wRfac Scor
S___1__1 1 106.45 50.40 1.37 0.352 0.000 0.273 10.20 0.595 0.390
S___2__1 2 0.00 0.00 0.00 0.352 0.000 0.273 11.44 0.601 0.377
S___4__1 3 22.21 -179.07 120.40 0.104 0.000 0.167 13.25 0.607 0.361
S___3_14 4 158.10 -1.79 120.96 0.489 0.000 0.386 5.00 0.614 0.353
S___5__1 5 0.83 144.00 179.87 0.216 0.000 0.272 3.76 0.614 0.349
S___7__1 6 0.00 0.00 179.92 0.216 0.000 0.272 3.77 0.617 0.342
S___8__6 7 34.01 88.19 71.45 0.178 0.000 0.186 2.12 0.622 0.332
S___6__6 8 146.73 90.95 73.37 0.368 0.000 0.225 0.27 0.625 0.325
S___9__3 9 77.12 174.71 90.84 0.482 0.000 0.245 2.06 0.626 0.318
S__10__2 10 134.42 132.29 7.11 0.075 0.000 0.233 1.50 0.627 0.317
S__11__2 11 152.93 177.57 136.00 0.091 0.000 0.216 2.94 0.628 0.316
S__13__6 12 130.28 116.07 92.63 0.473 0.000 0.197 1.20 0.629 0.315
S__12__5 13 156.31 -169.21 19.45 0.084 0.000 0.319 1.53 0.628 0.313
Contrast = 2.92
Sol_
S_ Nmon RF TF theta phi chi tx ty tz TFcnt wRfac Scor
S__ 2 1 1 106.45 50.40 1.37 0.352 0.000 0.273 10.20 0.595 0.390
PST_vector was used 1: 0.265 0.000 0.000
--- convert "molrep.crd" to "molrep.pdb" ---