Gene 5 protein

Summary of this structure solution:

This is a dataset with 3-wavelengths of MAD data, 2800 reflections to 2.6 A, 87 amino acids, and 2 selenium sites (Met-1, Met-77). SOLVE found both selenium sites in 34 minutes on an DEC alpha 500 MHz workstation. The Met-1 site has a very high thermal factor.

Solve.setup file listing basic information about the crystals:

CELL 76.08 27.97 42.36 90 103.2 90	! cell param
symfile /usr/local/lib/solve/c2.sym	! space group symmetry
resolution 2.6 20.0	! resolution limits

Input script file used to run SOLVE on gene 5 protein

#!/bin/csh

# set CCP4 and SOLVETMPDIR variables:

setenv CCP4_OPEN UNKNOWN

setenv SOLVETMPDIR /var/tmp

setenv SYMOP /usr/local/lib/solve/symop.lib

setenv SYMINFO /usr/local/lib/solve/syminfo.lib

solve <<EOD > solve.log

!command file to read in raw MAD data, scale, analyze and solve it----

checksolve ! compare solution with known h.a. sites

comparisonfile gvp.fft ! FFT map using FCALC from refined model

@solve.setup ! get our standard information read in

logfile mad.logfile ! write out most information to this file.

! summary info will be written to solve.prt

readformatted ! alternatives are readdenzo, readtrek

premerged ! alternative is unmerged

read_intensities ! alternative is read_amplitudes

refscattfactors ! alternative is fixscattfactors

mad_atomname se ! anomalously scattering atom is Se

lambda 1 ! info on wavelength #1 follows

label Wavelength # 1 ! a label for this wavelength

rawmadfile test_wva.fmt ! datafile with h k l Intensity sigma or

! h k l I+ sigma+ I- sigma-

wavelength 0.9000 ! wavelength value

fprimv_mad -1.6 ! f' value at this wavelength

fprprv_mad 3.4 ! f doubleprime value at this wavelength

! input refined h.a. coordinates (used only for comparison in "checksolve")

atomname se

XYZ 0.4813319 0.9972169 9.4140753E-02

XYZ 0.9731338 0.2875228 0.9446641

lambda 2

rawmadfile test_wvb.fmt

wavelength 0.9794

fprimv_mad -8.5

fprprv_mad 4.8

lambda 3

rawmadfile test_wvc.fmt

wavelength 0.9797

fprimv_mad -9.85

fprprv_mad 2.86

premerged

readformatted

nres 100 [approx # of residues in protein molecule]

nanomalous 2 [approx # of anomalously scattering atoms per protein]

SCALE_MAD ! read in and localscale the data

ANALYZE_MAD ! run MADMRG and MADBST and analyze all the Pattersons

SOLVE ! Solve the structure

EOD

Summary information from the "solve.prt" output file produced after completion of the automated structure determination

Correlation of anomalous differences. These indicate that the data beyond about 2.7 A are not contributing much to the phasing, as the correlation is less than 0.3.

            CORRELATION FOR

           WAVELENGTH PAIRS

 DMIN    1 VS 2   1 VS 3   2 VS 3

 5.20     0.90     0.72     0.80

 3.90     0.76     0.54     0.69

 3.64     0.69     0.44     0.69

 3.44     0.65     0.44     0.52

 3.25     0.59     0.27     0.50

 3.12     0.57     0.34     0.47

 2.99     0.58     0.18     0.39

 2.86     0.42     0.30     0.46

 2.73     0.33     0.17     0.31

 2.60     0.19     0.09     0.32

List of sites analyzed for compatibility with difference Patterson

   PEAK         X         Y         Z     OPTIMIZED

                                            RELATIVE OCCUPANCY        

     1     0.484     1.000     0.090      91.024        

     2     0.026     0.292     0.062      37.211     

Evaluation of this test soln with    2 sites after optimizing    

occupancy of each site     

Cross-vectors for sites  1 and  1 (excluding origin; 1000 = 1 sigma):    

#      U        V      W      HEIGHT   PRED HEIGHT      SYMM#

 1   -0.969   0.000  -0.181   16389.9     16570.9          2     





Cross-vectors for sites  2 and  1 (excluding origin; 1000 = 1 sigma):    

#      U        V      W      HEIGHT   PRED HEIGHT      SYMM#     

 1   -0.458  -0.708  -0.028   4053.98     3387.13          1     

 2   -0.510  -0.708  -0.153   4583.90     3387.13          1     





Cross-vectors for sites  2 and  2 (excluding origin; 1000 = 1 sigma):    

#      U        V      W      HEIGHT   PRED HEIGHT      SYMM#     

 1   -0.052   0.000  -0.125   1640.05     2769.35          2     





Overall quality of this Patterson soln =  7815.40       

Overall quality of the fit to patterson = 0.798126       

Avg normalized peak height =  3495.15

Selenium atom occupancy, coordinates, and thermal factors, and  

Cross-validation fouriers calculated with all heavy atoms in  

all derivs except the site being evaluated and any sites equivalent to it.

(Peak height is height of peak at this position/rms of map)      

Site    x       y       z       occ       B     -- PEAK  HEIGHT --         

  1   0.484   0.997   0.093   0.526  49.596              4.90      

  2   0.028   0.285   0.059   0.369  60.000              3.56

Re-refinement of f' and f" values:

                    Final refined values of f-prime and f"        

Wavelength  ------- f-prime --------       --------f"--------------          

     last refinement       Refined     last refinement       Refined            

1          0.327          0.327              3.817          2.928         

2         -8.419         -7.357              6.146          5.860         

3         -9.609         -8.668              2.272          1.774

Figure of merit versus resolution, and anomalous and dispersive FH/E versus resolution

 FIGURE OF MERIT WITH RESOLUTION   

DMIN:           TOTAL    8.81   5.75   4.55   3.88   3.44   3.12   2.88   2.68   

N:                2668    153    234    301    343    388    417    452    380   

MEAN FIG MERIT:   0.46   0.65   0.64   0.62   0.56   0.52   0.37   0.34   0.26

RMS ANOMALOUS FH/E  [f" PART OF FH / RMS ANO ERROR]:     



LAMBDA:  1         0.5    0.7    0.8    0.6    0.6    0.5    0.3    0.3    0.2   

LAMBDA:  2         0.6    0.8    0.8    0.7    0.7    0.6    0.5    0.4    0.3   

LAMBDA:  3         0.3    0.6    0.6    0.5    0.3    0.3    0.2    0.2    0.1     

RMS DISPERSIVE FH/E  [Delta-f-prime PART OF FH / RMS DISPERSIVE ERROR]:    



L1 VS L2:          0.8    1.0    1.1    1.0    0.9    0.8    0.5    0.4    0.3   

L1 VS L3:          0.9    1.2    1.3    1.0    1.0    0.9    0.6    0.5    0.4   

L2 VS L3:          0.2    0.4    0.4    0.3    0.2    0.2    0.1    0.1    0.1

The summary of scoring for this solution

 Summary of scoring for this solution:

                             -- over many solutions--    -- this solution --   

Criteria                       MEAN          SD         VALUE        Z-SCORE   

Pattersons:                   3.11         1.86         5.83         1.46   

Cross-validation Fourier:     3.26         2.44         7.05         1.55   

NatFourier CCx100:            27.4         5.39         31.3        0.729   

Mean figure of meritx100:    0.000E+00     8.26         58.5         7.08   

Correction for Z-scores:                                            -2.85     





Overall Z-score value:                                               7.97

Note that the Patterson and cross-validation Fourier scores are 1.5 sigma above the starting solutions, but native fourier analysis is just 1 sigma above. This is both because the asymmetric unit is small and the map is fairly noisy.

The end of the solve.status file:

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                      SOLVE STATUS      07-oct-00 10:34:53     

TIME ELAPSED:    34 MIN     

---------------------------------------------------------------------------   

CURRENT STEP:SOLVE MAIN PROGRAM   STATUS:   DONE   

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       ---TOP SOLUTION FOUND BY SOLVE  (<m> = 0.59; score =   8.03) ---

               X        Y        Z         OCCUP     B          HEIGHT/SIGMA       

        2     0.484    0.997    0.093     0.526     49.6              4.3

        2     0.028    0.285    0.059     0.369     60.0              4.4 

              TIME REQUIRED TO OBTAIN THIS SOLUTION:    28 MIN   

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CURRENT RESOLUTION:   2.6 A.    FINAL RESOLUTION:   2.6 A.

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