I was about to complain that I have 14+ domains/ASU, but then I started wondering—how on Earth did you do this sort of thing in the early days of crystallographic software/structure? No PISA back then, and I imagine things were far less automated…
JPK From: CCP4 bulletin board [mailto:[email protected]] On Behalf Of Eleanor Dodson Sent: Friday, February 06, 2015 6:00 AM To: [email protected] Subject: Re: [ccp4bb] Resolve Domain Sequence Ambiguities It looks a brilliant MR solution.. Do you know how you expect the A/B sets to pack? This was always a problem with heamoglobin where the A & B chains are very similar ad form a tetramer, but we would group the solutions iinto sensible tetramers (PISA will do that for you) and either set those residues which were different to ALA then try to use the density to check or fit the known teramer both ways so that one had the correct A/B match and the other did not, then let the refinement choose which fit was better. Eleanor On 5 February 2015 at 16:03, Dom Bellini <[email protected]<mailto:[email protected]>> wrote: Did you try ShelxE autotrace? At that resolution it should work nicely, or at least be able to distinguish between the two sequences. D -----Original Message----- From: CCP4 bulletin board [mailto:[email protected]<mailto:[email protected]>] On Behalf Of Keller, Jacob Sent: 05 February 2015 15:49 To: ccp4bb Subject: Re: [ccp4bb] Resolve Domain Sequence Ambiguities Sorry--should have included a bit more info: A huge caveat: data is tetartohedrally twinned, which makes everything an issue. But, I have managed to get Rfree down to ~32% in refmac with intensity-based twin refinement. I know--R's with de-twinning are illusory, but I still think it's solved, since I also can see anomalous signal in partial-model LLG maps from phaser at places predicted by the structure (and I don't think that anomalous info could be encoded in the MR phases (can it?)). So, that was a nice confirmatory moment... Resolution is good, but cut off by detector limit on home system (set at max resolution w/o 2theta) N 1/d^2 Dmid CCanom Nanom RCRanom CC1/2 NCC1/2 Rsplit CCfit CCanomfit $$ $$ 1 0.0061 12.82 -0.018 938 0.982 0.998 989 0.028 1.000 0.001 2 0.0182 7.40 0.026 1831 1.026 0.998 1880 0.027 1.000 0.006 3 0.0304 5.73 0.062 2334 1.064 0.995 2385 0.032 1.000 0.010 4 0.0426 4.85 0.048 2806 1.049 0.995 2881 0.033 1.000 0.014 5 0.0547 4.27 0.036 3164 1.036 0.995 3243 0.034 1.000 0.019 6 0.0669 3.87 0.025 3524 1.026 0.994 3606 0.035 1.000 0.023 7 0.0791 3.56 0.011 3758 1.011 0.993 3873 0.038 1.000 0.027 8 0.0912 3.31 0.021 4111 1.021 0.993 4206 0.041 0.999 0.032 9 0.1034 3.11 -0.003 4340 0.997 0.991 4441 0.044 0.999 0.036 10 0.1155 2.94 -0.040 4602 0.961 0.989 4724 0.049 0.999 0.040 11 0.1277 2.80 0.050 4864 1.051 0.991 4968 0.048 0.999 0.045 12 0.1399 2.67 0.051 4996 1.052 0.988 5121 0.052 0.998 0.049 13 0.1520 2.56 0.073 5348 1.076 0.988 5469 0.053 0.997 0.053 14 0.1642 2.47 -0.009 5486 0.991 0.987 5611 0.057 0.996 0.058 15 0.1764 2.38 0.032 5698 1.032 0.985 5827 0.061 0.995 0.062 16 0.1885 2.30 0.091 5945 1.095 0.985 6067 0.063 0.993 0.066 17 0.2007 2.23 0.038 6002 1.039 0.985 6118 0.064 0.991 0.070 18 0.2128 2.17 0.086 6331 1.089 0.983 6445 0.068 0.987 0.075 19 0.2250 2.11 0.075 6411 1.078 0.983 6556 0.070 0.983 0.079 20 0.2372 2.05 0.097 6608 1.102 0.983 6756 0.070 0.977 0.083 21 0.2493 2.00 0.141 6712 1.152 0.981 6880 0.078 0.968 0.088 22 0.2615 1.96 0.104 6556 1.110 0.974 7126 0.093 0.957 0.092 23 0.2737 1.91 0.141 5595 1.152 0.953 7054 0.121 0.943 0.096 24 0.2858 1.87 0.149 4659 1.160 0.936 6652 0.151 0.923 0.101 25 0.2980 1.83 0.125 3630 1.133 0.909 5936 0.184 0.898 0.105 26 0.3101 1.80 0.138 2640 1.148 0.874 5118 0.219 0.866 0.109 27 0.3223 1.76 0.111 1793 1.116 0.851 4035 0.250 0.825 0.114 28 0.3345 1.73 0.040 1040 1.041 0.782 2857 0.305 0.776 0.118 29 0.3466 1.70 0.094 300 1.097 0.713 1474 0.371 0.717 0.122 30 0.3588 1.67 - 2 - 0.589 228 0.497 0.649 0.127 $$ Overall: 0.027 122024 1.027 0.997 138526 0.048 CCanom Nanom RCRanom CC1/2 NCC1/2 Rsplit CCfit CCanomfit MR solution: SOLU SET RFZ=3.9 TFZ=7.1 PAK=0 LLG=38 RFZ=3.7 TFZ=8.9 PAK=0 LLG=75 RFZ=4.2 TFZ=8.2 PAK=0 LLG=40 LLG=160 RFZ=3.2 TFZ=11.2 PAK=0 LLG=249 LLG=262 RFZ=4.2 TFZ=9.4 PAK=0 LLG=331 LLG=339 RFZ=4.1 TFZ=11.3 PAK=0 LLG=405 LLG=418 RFZ=3.9 TFZ=11.6 PAK=0 LLG=487 LLG=494 RFZ=4.2 TFZ=8.9 PAK=0 LLG=539 LLG=554 RFZ=3.3 TFZ=8.6 PAK=0 LLG=612 LLG=638 RFZ=3.1 TFZ=8.7 PAK=0 LLG=679 LLG=691 RFZ=2.6 TFZ=12.4 PAK=0 LLG=910 LLG=926 RFZ=3.0 TFZ=9.2 PAK=0 LLG=987 LLG=1001 RFZ=2.7 TFZ=9.8 PAK=3 LLG=1018 LLG=1043 RFZ=2.9 TFZ=9.1 PAK=5 LLG=1039 -----Original Message----- From: [email protected]<mailto:[email protected]> [mailto:[email protected]<mailto:[email protected]>] Sent: Thursday, February 05, 2015 10:28 AM To: Keller, Jacob; [email protected]<mailto:[email protected]> Subject: RE: Resolve Domain Sequence Ambiguities Dear Jacob, Are you sure your MR solution is correct? What is your resolution? Best, D -----Original Message----- From: CCP4 bulletin board [mailto:[email protected]<mailto:[email protected]>] On Behalf Of Keller, Jacob Sent: 05 February 2015 15:21 To: ccp4bb Subject: [ccp4bb] Resolve Domain Sequence Ambiguities Hi All, I have 14 identical domains placed in my ASU with reasonable MR scores, but the problem is that there should really be 7+7 or 8+8 of domains A and B (which are structurally similar). Can anyone think of a great and easy way of resolving the ambiguity? I was thinking potentially: -change one domain to polyAla -SA omit map of that one -rebuild/refine -iterate through all domains, noting scores Seems it might be pretty low reliability and a fair amount of work though. Otherwise, could try to go back to a small SAD signal, use partial model phases to find HAs, then phase without the model, rebuild from there. Any thoughts or similar experiences? JPK ******************************************* Jacob Pearson Keller, PhD Looger Lab/HHMI Janelia Research Campus 19700 Helix Dr, Ashburn, VA 20147 email: [email protected]<mailto:[email protected]> ******************************************* -- This e-mail and any attachments may contain confidential, copyright and or privileged material, and are for the use of the intended addressee only. If you are not the intended addressee or an authorised recipient of the addressee please notify us of receipt by returning the e-mail and do not use, copy, retain, distribute or disclose the information in or attached to the e-mail. Any opinions expressed within this e-mail are those of the individual and not necessarily of Diamond Light Source Ltd. Diamond Light Source Ltd. cannot guarantee that this e-mail or any attachments are free from viruses and we cannot accept liability for any damage which you may sustain as a result of software viruses which may be transmitted in or with the message. 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