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CCP4i: Graphical User Interface |
| MIR Tutorial Bath - MLPHARE / VECREF |
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8. Suggested procedure for phasing and refinement with MLPHARE / VECREF
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Compute the isomorphous difference Pattersons for all derivatives to the maximum resolution of the derivative (i.e. without resolution cutoffs).
# fft HKLIN uffhscal MAPOUT ufdp <<EOD TITLE UF FH scaled native 3A isodif Patterson. PATT BIAS 1 ! Compensate for over-estimation of weak reflections. LABIN F1=FUF SIG1=SIGFUF F2=FNAT SIG2=SIGFNAT GRID 120 108 72 VF000 151447.56 ! "Volume" from FHSCAL output. EOD if ($status) exit fft HKLIN uafhscal MAPOUT uadp <<EOD TITLE UAc FH scaled native 3A isodif Patterson. PATT BIAS 1 LABIN F1=FUAC SIG1=SIGFUAC F2=FNAT SIG2=SIGFNAT GRID 120 108 72 VF000 151595.69 ! "Volume" from FHSCAL output. EOD if ($status) exit fft HKLIN usfhscal MAPOUT usdp <<EOD TITLE US FH scaled native 3A isodif Patterson. PATT BIAS 1 LABIN F1=FUS SIG1=SIGFUS F2=FNAT SIG2=SIGFNAT GRID 120 108 72 VF000 133105.75 ! "Volume" from FHSCAL output. EOD if ($status) exit fft HKLIN ptfhscal MAPOUT ptdp <<EOD TITLE PtCl FH scaled native 3A isodif Patterson. PATT BIAS 1 LABIN F1=FPTCL SIG1=SIGFPTCL F2=FNAT SIG2=SIGFNAT GRID 120 108 72 VF000 146681.38 ! "Volume" from FHSCAL output. EOD if ($status) exit fft HKLIN ybfhscal MAPOUT ybdp <<EOD TITLE YbCl FH scaled native 3A isodif Patterson. PATT BIAS 1 LABIN F1=FYBCL SIG1=SIGFYBCL F2=FNAT SIG2=SIGFNAT GRID 120 108 72 VF000 160414.22 ! "Volume" from FHSCAL output. EOD if ($status) exit fft HKLIN hgfhscal MAPOUT hgdp <<EOD TITLE PCMBS FH scaled native 3A isodif Patterson. PATT BIAS 1 LABIN F1=FPCMBS SIG1=SIGFPCMBS F2=FNAT SIG2=SIGFNAT GRID 120 108 72 VF000 150653.00 ! "Volume" from FHSCAL output. EOD
Again, note that the BIAS factor, whose function is to reduce the contribution to the Patterson of the weak high resolution reflections, which would otherwise contribute mainly noise, should only be used if the standard deviations of the amplitudes are reliable.
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Provided there is no evidence of non-isomorphism, from the "best" Patterson, locate 1 or 2 major sites.
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Refine with VECREF the site occupancies alone, then the occupancies and coordinates, then these plus the B parameters (this is all done in one job). Note that the site coordinates have been modified from those produced by the run of VECSUM (section 4.3.4.) with an origin shift of (0,½,0), together with a symmetry transformation of the space group. This was done purely to allow easy comparison of the resultant phases with those produced by the previous MLPHARE procedure.
# vecref MAPIN ufdp ATOUT ufvr.xyz <<EOD TITLE VECREF PBGD UF deriv - native. Isodif 3A. SPACEG P21212 RESOL 20 3 CYCLES 3 5 5 BREF ATOM U 1 1 .28 .18 .84 25 ATOM U 2 1 .68 .18 .34 25 EOD
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Using MLPHARE in phase-only mode, calculate phases based on this derivative.
# mlphare HKLIN pbgd_fhscal HKLOUT ufmp <<EOD CYCLES 3 LABIN FP=FNAT SIGFP=SIGFNAT - FPH1=FUF SIGFPH1=SIGFUF LABOUT ALLIN PRINT AVF AVE DERIV UF DCYCLE PHASE ALL ATOM U 0.2864 0.1706 0.8346 1.0197 1.0197 BFAC 29.45 ATOM U 0.6840 0.1865 0.3449 0.6344 0.6344 BFAC 45.14 EOD -
Using FFT and PEAKMAX, get all the peaks in the difference Fourier higher than a specified threshold (4s is suggested), with symmetry-related peaks eliminated. This can either be done by manually editing the logfile output from PEAKMAX, or writing a jiffy program to extract the sites, check for symmetry duplicates, and reformat the site coordinates ready for insertion into the VECREF script. The occupancies can be set all equal to 1, or better still, scaled in proportion to the peak heights; the B factors can be initialised to 25.
# fft HKLIN ufmp MAPOUT ufmpdf <<EOD TITLE UF diff Fourier phased on UF derivative refined by VECREF. LABIN F1=FUF SIG1=SIGFUF F2=FNAT SIG2=SIGFNAT PHI=PHIB W=FOM EOD if ($status) exit peakmax MAPIN ufmpdf <<EOD THRESH RMS 4 OUTPUT NONE EOD rm ufmpdf.map
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Put all sites in together to refine with VECREF, as in step m.
The program will eliminate sites that either refine to low occupancy (e.g. < 2s), or to unrealistically low (e.g. < 0) or high (e.g. > 200) values of the B parameter.
# vecref MAPIN ufdp ATOUT ufvr1.xyz <<EOD TITLE VECREF PBGD UF deriv - native. Isodif 3A. SPACEG P21212 RESOL 20 3 CYCLES 3 5 10 BREF ATOM U 1 1 .28615 0.17076 0.83455 25 ATOM U 2 1 .68374 0.18610 0.34608 25 ATOM U 3 1 .18484 0.19094 0.89099 25 ATOM U 4 1 .28470 0.23418 0.83511 25 ATOM U 5 1 .28609 0.10676 0.83399 25 ATOM U 6 1 .28079 0.16491 0.94649 25 ATOM U 7 1 .50025 0.25000 0.61053 25 ATOM U 8 1 .28066 0.16519 0.72478 25 ATOM U 9 1 .78356 0.16182 0.40031 25 EOD
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Repeat steps n and o, but calculate the cross-difference Fourier for the next derivative.
# mlphare HKLIN pbgd_fhscal HKLOUT ufmp <<EOD CYCLES 3 LABIN FP=FNAT SIGFP=SIGFNAT - FPH1=FUF SIGFPH1=SIGFUF LABOUT ALLIN PRINT AVF AVE DERIV UF DCYCLE PHASE ALL ATOM U 0.2859 0.1691 0.8332 1.4204 1.4204 BFAC 34.96 ATOM U 0.6825 0.1874 0.3440 0.5731 0.5731 BFAC 46.02 ATOM U 0.1846 0.1869 0.8877 0.4607 0.4607 BFAC 11.85 ATOM U 0.5005 0.2449 0.6131 0.4474 0.4474 BFAC 17.45 EOD if ($status) exit fft HKLIN ufmp MAPOUT uampdf <<EOD TITLE UAc diff Fourier phased on UF derivative refined by VECREF. LABIN F1=FUAC SIG1=SIGFUAC F2=FNAT SIG2=SIGFNAT PHI=PHIB W=FOM EOD if ($status) exit peakmax MAPIN uampdf <<EOD THRESH RMS 4 OUTPUT NONE EOD rm ufmp.mtz uampdf.map -
Refine all sites together for this derivative, as in step p.
# vecref MAPIN uadp ATOUT uavr1.xyz <<EOD TITLE VECREF PBGD UAc deriv - native. Isodif 3A. SPACEG P21212 RESOL 20 3 CYCLES 3 5 10 BREF ATOM U 1 1 .28561 0.16973 0.83464 25 ATOM U 2 1 .18535 0.18616 0.88867 25 ATOM U 3 1 .49905 0.25000 0.61358 25 ATOM U 4 1 .68243 0.18754 0.34529 25 ATOM U 5 1 .28067 0.10440 0.83276 25 ATOM U 6 1 .04065 0.25000 0.46040 25 ATOM U 7 1 .28534 0.23489 0.82928 25 EOD
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Add all the new refined sites for the new derivative to the input for MLPHARE.
Repeat steps q and r for each of the remaining derivatives.
# mlphare HKLIN pbgd_fhscal HKLOUT uamp <<EOD CYCLES 3 LABIN FP=FNAT SIGFP=SIGFNAT - FPH1=FUF SIGFPH1=SIGFUF - FPH2=FUAC SIGFPH2=SIGFUAC LABOUT ALLIN PRINT AVF AVE DERIV UF DCYCLE PHASE ALL ATOM U 0.2859 0.1691 0.8332 1.4204 1.4204 BFAC 34.96 ATOM U 0.6825 0.1874 0.3440 0.5731 0.5731 BFAC 46.02 ATOM U 0.1846 0.1869 0.8877 0.4607 0.4607 BFAC 11.85 ATOM U 0.5005 0.2449 0.6131 0.4474 0.4474 BFAC 17.45 DERIV UAc DCYCLE PHASE ALL ATOM U 0.2881 0.1705 0.8453 0.7896 0.7896 BFAC 41.58 ATOM U 0.1844 0.1846 0.8904 0.9079 0.9079 BFAC 16.16 ATOM U 0.4855 0.2383 0.6310 0.5954 0.5954 BFAC 22.73 ATOM U 0.6819 0.1883 0.3428 0.2363 0.2363 BFAC 37.83 ATOM U 0.0398 0.2416 0.4586 0.7912 0.7912 BFAC 45.86 EOD if ($status) exit fft HKLIN uamp MAPOUT usmpdf <<EOD TITLE US diff Fourier phased on UF & UAc derivatives refined by VECREF. LABIN F1=FUS SIG1=SIGFUS F2=FNAT SIG2=SIGFNAT PHI=PHIB W=FOM EOD if ($status) exit peakmax MAPIN usmpdf <<EOD THRESH RMS 4 OUTPUT NONE EOD rm uamp.mtz usmpdf.map # vecref MAPIN usdp ATOUT usvr1.xyz <<EOD TITLE VECREF PBGD US deriv - native. Isodif 3A. SPACEG P21212 RESOL 20 3 CYCLES 3 5 10 BREF ATOM U 1 1 .18184 0.19281 0.88767 25 ATOM U 1 1 .28535 0.17936 0.83471 25 ATOM U 1 1 .66149 0.13307 0.90081 25 ATOM U 1 1 .69055 0.17053 0.85079 25 ATOM U 1 1 .87630 0.05247 0.65764 25 ATOM U 1 1 .49026 0.23516 0.63313 25 ATOM U 1 1 .71442 0.13248 0.76857 25 ATOM U 1 1 .67905 0.03676 0.10827 25 ATOM U 1 1 .89558 0.14894 0.14228 25 ATOM U 1 1 .78685 0.17541 0.81784 25 EOD # mlphare HKLIN pbgd_fhscal HKLOUT usmp <<EOD CYCLES 3 LABIN FP=FNAT SIGFP=SIGFNAT - FPH1=FUF SIGFPH1=SIGFUF - FPH2=FUAC SIGFPH2=SIGFUAC - FPH3=FUS SIGFPH3=SIGFUS LABOUT ALLIN PRINT AVF AVE DERIV UF DCYCLE PHASE ALL ATOM U 0.2859 0.1691 0.8332 1.4204 1.4204 BFAC 34.96 ATOM U 0.6825 0.1874 0.3440 0.5731 0.5731 BFAC 46.02 ATOM U 0.1846 0.1869 0.8877 0.4607 0.4607 BFAC 11.85 ATOM U 0.5005 0.2449 0.6131 0.4474 0.4474 BFAC 17.45 DERIV UAc DCYCLE PHASE ALL ATOM U 0.2881 0.1705 0.8453 0.7896 0.7896 BFAC 41.58 ATOM U 0.1844 0.1846 0.8904 0.9079 0.9079 BFAC 16.16 ATOM U 0.4855 0.2383 0.6310 0.5954 0.5954 BFAC 22.73 ATOM U 0.6819 0.1883 0.3428 0.2363 0.2363 BFAC 37.83 ATOM U 0.0398 0.2416 0.4586 0.7912 0.7912 BFAC 45.86 DERIV US DCYCLE PHASE ALL ATOM U 0.1826 0.1927 0.8884 0.7042 0.7042 BFAC 30.66 EOD if ($status) exit fft HKLIN usmp MAPOUT ptmpdf <<EOD TITLE PtCl diff Fourier phased on UF, UAc & US derivatives refined by VECREF. LABIN F1=FPTCL SIG1=SIGFPTCL F2=FNAT SIG2=SIGFNAT PHI=PHIB W=FOM EOD if ($status) exit peakmax MAPIN ptmpdf <<EOD THRESH RMS 4 OUTPUT NONE EOD rm usmp.mtz ptmpdf.map # vecref MAPIN ptdp ATOUT ptvr1.xyz <<EOD TITLE VECREF PBGD PtCl deriv - native. Isodif 3A. SPACEG P21212 RESOL 20 3 CYCLES 3 5 10 BREF ATOM PT 1 1 .25247 0.04321 0.40324 25 ATOM PT 2 1 .17795 0.20318 0.89739 25 ATOM PT 3 1 .15464 0.04206 0.53930 25 ATOM PT 4 1 .28551 0.13117 0.84088 25 ATOM PT 5 1 .28334 0.20094 0.83942 25 ATOM PT 6 1 .18329 0.15558 0.91884 25 ATOM PT 7 1 .08052 0.23379 0.40924 25 ATOM PT 8 1 .12630 0.15489 0.41827 25 ATOM PT 9 1 .28262 0.16810 0.76787 25 ATOM PT 10 1 .00000 0.00000 0.53845 25 EOD # mlphare HKLIN pbgd_fhscal HKLOUT ptmp <<EOD CYCLES 3 LABIN FP=FNAT SIGFP=SIGFNAT - FPH1=FUF SIGFPH1=SIGFUF - FPH2=FUAC SIGFPH2=SIGFUAC - FPH3=FUS SIGFPH3=SIGFUS - FPH4=FPTCL SIGFPH4=SIGFPTCL LABOUT ALLIN PRINT AVF AVE DERIV UF DCYCLE PHASE ALL ATOM U 0.2859 0.1691 0.8332 1.4204 1.4204 BFAC 34.96 ATOM U 0.6825 0.1874 0.3440 0.5731 0.5731 BFAC 46.02 ATOM U 0.1846 0.1869 0.8877 0.4607 0.4607 BFAC 11.85 ATOM U 0.5005 0.2449 0.6131 0.4474 0.4474 BFAC 17.45 DERIV UAc DCYCLE PHASE ALL ATOM U 0.2881 0.1705 0.8453 0.7896 0.7896 BFAC 41.58 ATOM U 0.1844 0.1846 0.8904 0.9079 0.9079 BFAC 16.16 ATOM U 0.4855 0.2383 0.6310 0.5954 0.5954 BFAC 22.73 ATOM U 0.6819 0.1883 0.3428 0.2363 0.2363 BFAC 37.83 ATOM U 0.0398 0.2416 0.4586 0.7912 0.7912 BFAC 45.86 DERIV US DCYCLE PHASE ALL ATOM U 0.1826 0.1927 0.8884 0.7042 0.7042 BFAC 30.66 DERIV PtCl DCYCLE PHASE ALL ATOM PT 0.2507 0.0452 0.4118 1.2751 1.2751 BFAC 49.26 ATOM PT 0.1676 0.1937 0.9093 0.4910 0.4910 BFAC 96.21 ATOM PT 0.1568 0.0414 0.5378 0.7443 0.7443 BFAC 55.40 ATOM PT 0.1490 0.1411 0.9022 0.4471 0.4471 BFAC 74.56 ATOM PT 0.0770 0.2304 0.4085 0.3281 0.3281 BFAC 50.08 ATOM PT 0.1217 0.1374 0.4133 0.3520 0.3520 BFAC 19.55 EOD if ($status) exit fft HKLIN ptmp MAPOUT ybmpdf <<EOD TITLE YbCl diff Fourier phased on UF, UAc, US & Pt derivatives. LABIN F1=FYBCL SIG1=SIGFYBCL F2=FNAT SIG2=SIGFNAT PHI=PHIB W=FOM EOD if ($status) exit peakmax MAPIN ybmpdf <<EOD THRESH RMS 4 OUTPUT NONE EOD rm ptmp.mtz ybmpdf.map # vecref MAPIN ybdp ATOUT ybvr1.xyz <<EOD TITLE VECREF PBGD YbCl deriv - native. Isodif 3A. SPACEG P21212 RESOL 20 3 CYCLES 3 5 10 BREF ATOM YB 1 1 .49373 0.23695 0.61978 25 ATOM YB 2 1 .18322 0.18647 0.88976 25 ATOM YB 3 1 .28605 0.16971 0.83379 25 ATOM YB 4 1 .98864 0.25000 0.38272 25 ATOM YB 5 1 .00927 0.05067 0.47647 25 ATOM YB 6 1 .87544 0.03871 0.63246 25 ATOM YB 7 1 .95501 0.13334 0.16714 25 ATOM YB 8 1 .24534 0.06416 0.15415 25 ATOM YB 9 1 .68375 0.18664 0.33213 25 EOD -
Use MLPHARE in phased refinement mode with the anomalous data to detect the correct hand, as previously described.
If necessary, correct the input coordinates and set the anomalous occupancy of each atom to its real occupancy.
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Using MLPHARE in phasing-only mode, calculate phases based on all derivatives (again, the PCMBS derivative was omitted).
# mlphare HKLIN pbgd_fhscal HKLOUT allmp1 <<EOD CYCLES 3 LABIN FP=FNAT SIGFP=SIGFNAT - FPH1=FUF SIGFPH1=SIGFUF - FPH2=FUAC SIGFPH2=SIGFUAC - FPH3=FUS SIGFPH3=SIGFUS - FPH4=FPTCL SIGFPH4=SIGFPTCL - FPH5=FYBCL SIGFPH5=SIGFYBCL LABOUT ALLIN PRINT AVF AVE DERIV UF DCYCLE PHASE ALL ATOM U 0.2859 0.1691 0.8332 1.4204 1.4204 BFAC 34.96 ATOM U 0.6825 0.1874 0.3440 0.5731 0.5731 BFAC 46.02 ATOM U 0.1846 0.1869 0.8877 0.4607 0.4607 BFAC 11.85 ATOM U 0.5005 0.2449 0.6131 0.4474 0.4474 BFAC 17.45 DERIV UAc DCYCLE PHASE ALL ATOM U 0.2881 0.1705 0.8453 0.7896 0.7896 BFAC 41.58 ATOM U 0.1844 0.1846 0.8904 0.9079 0.9079 BFAC 16.16 ATOM U 0.4855 0.2383 0.6310 0.5954 0.5954 BFAC 22.73 ATOM U 0.6819 0.1883 0.3428 0.2363 0.2363 BFAC 37.83 ATOM U 0.0398 0.2416 0.4586 0.7912 0.7912 BFAC 45.86 DERIV US DCYCLE PHASE ALL ATOM U 0.1826 0.1927 0.8884 0.7042 0.7042 BFAC 30.66 DERIV PtCl DCYCLE PHASE ALL ATOM PT 0.2507 0.0452 0.4118 1.2751 1.2751 BFAC 49.26 ATOM PT 0.1676 0.1937 0.9093 0.4910 0.4910 BFAC 96.21 ATOM PT 0.1568 0.0414 0.5378 0.7443 0.7443 BFAC 55.40 ATOM PT 0.1490 0.1411 0.9022 0.4471 0.4471 BFAC 74.56 ATOM PT 0.0770 0.2304 0.4085 0.3281 0.3281 BFAC 50.08 ATOM PT 0.1217 0.1374 0.4133 0.3520 0.3520 BFAC 19.55 DERIV YbCl DCYCLE PHASE ALL ATOM YB 0.4938 0.2410 0.6152 0.8800 0.8800 BFAC 33.38 EOD -
Repeat all the difference Fouriers and the VECREF refinements as in steps o and p.
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Repeat steps u and v until a consistent set of sites is attained (3 iterations).
# mlphare HKLIN pbgd_fhscal HKLOUT allmp4 <<EOD CYCLES 5 LABIN FP=FNAT SIGFP=SIGFNAT - FPH1=FUF SIGFPH1=SIGFUF DPH1=DANUF SIGDPH1=SIGDANUF - FPH2=FUAC SIGFPH2=SIGFUAC DPH2=DANUAC SIGDPH2=SIGDANUAC - FPH3=FUS SIGFPH3=SIGFUS - FPH4=FPTCL SIGFPH4=SIGFPTCL DPH4=DANPTCL SIGDPH4=SIGDANPTCL - FPH5=FYBCL SIGFPH5=SIGFYBCL - FPH6=FPCMBS SIGFPH6=SIGFPCMBS DPH6=DANPCMBS SIGDPH6=SIGDANPCMBS LABOUT ALLIN PRINT AVF AVE DERIV UF DCYCLE PHASE ALL ATOM U 0.2857 0.1706 0.8364 1.0553 1.0553 BFAC 31.71 ATOM U 0.1843 0.1875 0.8893 0.5338 0.5338 BFAC 19.32 ATOM U 0.5004 0.2448 0.6133 0.4753 0.4753 BFAC 17.22 ATOM U 0.6822 0.1878 0.3420 0.6059 0.6059 BFAC 51.09 ATOM U 0.9892 0.0538 0.4883 0.2830 0.2830 BFAC 29.83 DERIV UAc DCYCLE PHASE ALL ATOM U 0.1848 0.1849 0.8905 0.9219 0.9219 BFAC 17.69 ATOM U 0.2888 0.1707 0.8472 0.6473 0.6473 BFAC 26.28 ATOM U 0.4857 0.2386 0.6317 0.6174 0.6174 BFAC 21.81 ATOM U 0.0400 0.2422 0.4582 0.8310 0.8310 BFAC 49.24 ATOM U 0.6796 0.1902 0.3431 0.2445 0.2445 BFAC 35.64 ATOM U 0.9832 0.0503 0.4903 0.4344 0.4344 BFAC 45.67 ATOM U 0.9691 0.0262 0.1006 0.3802 0.3802 BFAC 61.76 DERIV US DCYCLE PHASE ALL ATOM U 0.1826 0.1933 0.8889 0.7029 0.7029 BFAC 29.99 DERIV PtCl DCYCLE PHASE ALL ATOM PT 0.2507 0.0452 0.4117 1.2707 1.2707 BFAC 49.47 ATOM PT 0.1569 0.0412 0.5374 0.7500 0.7500 BFAC 55.47 ATOM PT 0.1216 0.1371 0.4125 0.3743 0.3743 BFAC 26.08 ATOM PT 0.1649 0.1971 0.9063 0.4686 0.4686 BFAC 81.33 ATOM PT 0.0769 0.2298 0.4087 0.3164 0.3164 BFAC 49.01 ATOM PT 0.1509 0.1408 0.9052 0.4554 0.4554 BFAC 72.34 DERIV YbCl DCYCLE PHASE ALL ATOM YB 0.4976 0.2364 0.6143 0.5937 0.5937 BFAC 36.75 ATOM YB 0.1983 0.1838 0.9253 0.1541 0.1541 BFAC 26.52 ATOM YB 0.0106 0.0529 0.4774 0.1843 0.1843 BFAC 21.92 DERIV PCMBS DCYCLE PHASE ALL ATOM HG 0.0723 0.0646 0.1533 0.3838 0.3838 BFAC 35.06 ATOM HG 0.2452 0.0609 0.1442 0.2874 0.2874 BFAC 31.25 ATOM HG 0.7974 0.1644 0.7249 0.2557 0.2557 BFAC 63.77 EOD
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