Dear Perrakis,
The R merge of the lowest resolution shell is 0.08.
The matthews' analysisi is below:
For estimated molecular weight 78750.
Nmol/asym Matthews Coeff %solvent P(2.90) P(tot)
_______________________________
1 7.90 84.45 0.00 0.00
2 3.95 68.89 0.06 0.04
3 2.63 53.34 0.69 0.62
4 1.98 37.78 0.25 0.33
5 1.58 22.23 0.00 0.00
6 1.32 6.67 0.00 0.00
Do you mean that it is possible that there is only 2 molecules in the ASU?
I have also performed a 4.5 SF with Molrep. The log file is pasted as below:
Number of RF peaks : 10
theta phi chi alpha beta gamma Rf
Rf/sigma
Sol_RF 1 0.00 0.00 0.00 0.00 0.00 0.00 0.1101E+06
10.89
Sol_RF 2 0.00 0.00 30.00 30.00 0.00 0.00 0.3374E+05
3.34
Sol_RF 3 90.00 -68.59 179.99 0.00 180.00 -42.83 0.2630E+05
2.60
Sol_RF 4 148.78 30.00 179.98 30.00 -62.44 150.00 7709.
0.76
Sol_RF 5 117.57 30.00 179.99 30.00 -124.86 150.00 6042.
0.60
Sol_RF 6 56.28 60.00 179.98 60.00 -112.56 120.00 5925.
0.59
Sol_RF 7 90.00 -60.00 90.00 30.00 -90.00 -30.00 5917.
0.59
Sol_RF 8 90.00 -30.00 60.94 60.00 -60.94 -60.00 5485.
0.54
Sol_RF 9 135.89 -165.91 179.98 14.09 88.22 165.91 5424.
0.54
Sol_RF 10 138.31 -169.98 179.99 10.02 83.38 169.98 5249.
0.52
I think the SF function indicate that there are three molecules in the ASU.
Am i right?
Thansk and best wishes.
On Wed, Jul 15, 2009 at 2:27 PM, Anastassis Perrakis <a.perra...@nki.nl>wrote:
> Hi all -
> On 15 Jul 2009, at 7:03, Lijun Liu wrote:
>
> Hello dear Wei,
>
> 1) Your dataset has a high overall Rmerge. The outmost shell (70%) is very
> high, which suggests a need to shrink resolution. What about I/s(I),
> redundancy and completeness? Also, how many reflections (percentage) have
> been subjected to rejection? Too many rejections may mean a SG error.
>
>
> 70% Rmerge alone in the outer shell, can indeed be reasonable for such high
> symmetry, depending on e.g. redundancy and what Lijun also mentions. I would
> be more interested in the low resolution Rmerge to be able to have an
> opinion about the dataset ...
>
>
> 2) Software analyses including Phenix.xtriage give good suggestions, but
> they should be used reasonably. P6n22 could be twinned from P6n or P3n12 or
> P3n21 or even P3n, when close to be perfectly twinned.
>
>
> Correct, but phenix.xtriage as well as Pointless, will still pick up
> possible twining even if you have merged data to P6n. Still, its best to
> integrate the data to P1, which is always perfectly legitimate, and then run
> one of these programs to get the likelihood of the correct space group,
> and consider twining.
>
> 3) Please try a little more analyses of the cell shape and A.U. shape (very
> long in your case), with Matthews' coefficient analyses, you may get more
> hints about packing and SG.
>
>
> Indeed, for example keep in mind that for 2.9 A resolution from a good
> beamline, you might have a solvent content of 70%, quite easily.
>
> 4) Try to find one molecule first in such a big A.U. [Automatic scripts
> have a higher chance to fail than a smart manual scrip (very personal)].
>
>
> Here I do disagree, since I found that automated scripts
> are explicitly more useful in the case of multiple copies, and smarter than
> (my) old manual scripts. The reason is that they are likely to keep a lot of
> initial solutions for the first molecule, and then look for the second based
> on these many possibilities for the first, and so on, which can be very
> powerful.
>
> 5) Use a lower resolution ~4-5 Å data to do a better self rotation
> function. Why do not paste a .ps plot for the SR peaks.
>
>
> Using low resolution for SF is a good idea, 5-6 is also nice. But please do
> not send us any attachments at the BB ;-)
>
> A.
>
>
>
> Good luck.
>
> Lijun
>
> On Jul 14, 2009, at 4:23 AM, Wei Zhang wrote:
>
> Dears,
> I am doing molecular replacement of a protein complex with a P622 data set
> with large cell parameters (a=b=135, c=480). The data set seems well. R
> merge is 0.17 for all and 0.70 for the last shell of 2.9 angstrom. I am not
> sure it is a complex in the crystal. Phenix analysis reveal there is no
> twin. The proposed protein complex is about 70 kDa with a larger subunit of
> 50 kDa and a small subunit of 20 kDa. The matthews analysis indicates that
> there might be 3 complexes in the ASU. The structure of the 50 kDa subunit
> is known while the 20 kDa one is unknown. But molecular replacement failed
> with either Phaser or Molrep.
> Self-rotation with CNS reported the result as below:
> ! index, psi, phi, kappa, RF-function ( 0.25)
> 1 0.000 0.000 180.000 29.7217
> Self-ratation with Molrep reported the result as below:
> Number of RF peaks : 30
> theta phi chi alpha beta gamma Rf
> Rf/sigma
> Sol_RF 1 0.00 0.00 0.00 0.00 0.00 0.00 0.3444E+05
> 17.13
> Sol_RF 2 90.00 -80.07 179.99 0.00 180.00 -19.86 5061.
> 2.52
> Sol_RF 3 90.00 -65.53 179.99 0.00 180.00 -48.94 4890.
> 2.43
> Sol_RF 4 90.00 -76.12 179.99 0.00 180.00 -27.76 4722.
> 2.35
> Sol_RF 5 11.42 90.00 61.00 30.00 11.53 30.00 2438.
> 1.21
> Sol_RF 6 164.38 60.00 179.99 60.00 -31.24 120.00 1850.
> 0.92
> Sol_RF 7 85.13 -141.54 179.97 38.32 -170.26 141.40 1805.
> 0.90
> Sol_RF 8 90.00 -60.00 90.00 30.00 -90.00 -30.00 1764.
> 0.88
> Sol_RF 9 84.07 -144.87 179.96 34.98 -168.15 144.73 1743.
> 0.87
> Sol_RF 10 72.25 -60.00 89.30 46.76 -84.03 -13.24 1665.
> 0.83
> Sol_RF 11 138.24 -130.77 180.00 49.23 83.52 130.77 1608.
> 0.80
> Sol_RF 12 170.36 30.00 179.99 30.00 -19.28 150.00 1590.
> 0.79
> Sol_RF 13 82.04 60.00 179.99 60.00 -164.07 120.00 1571.
> 0.78
> Sol_RF 14 141.01 30.00 179.99 30.00 -77.99 150.00 1554.
> 0.77
> Sol_RF 15 123.63 -155.38 179.98 24.62 112.74 155.38 1517.
> 0.75
> Sol_RF 16 148.80 30.00 179.99 30.00 -62.39 150.00 1450.
> 0.72
> Sol_RF 17 142.61 30.00 180.00 30.00 -74.78 150.00 1439.
> 0.72
> Sol_RF 18 72.25 -150.00 179.99 30.00 -144.50 150.00 1422.
> 0.71
> Sol_RF 19 47.42 -120.00 179.98 60.00 -94.83 120.00 1417.
> 0.71
> Sol_RF 20 153.12 30.00 179.98 30.00 -53.75 150.00 1292.
> 0.64
> Sol_RF 21 155.99 30.00 179.98 30.00 -48.01 150.00 1274.
> 0.63
> Sol_RF 22 138.05 30.00 179.99 30.00 -83.91 150.00 1258.
> 0.63
> Sol_RF 23 54.12 120.00 89.15 60.00 69.32 0.00 1254.
> 0.62
> Sol_RF 24 129.04 -136.48 179.99 43.52 101.93 136.48 1193.
> 0.59
> Sol_RF 25 138.65 -164.29 179.99 15.71 82.69 164.29 1185.
> 0.59
> Sol_RF 26 84.16 51.37 179.98 51.37 -168.32 128.63 1176.
> 0.59
> Sol_RF 27 90.00 150.00 59.82 60.00 59.82 -60.00 1162.
> 0.58
> Sol_RF 28 43.66 -133.74 179.98 46.26 -87.31 133.74 1161.
> 0.58
> Sol_RF 29 127.85 -169.33 179.99 10.67 104.29 169.33 1153.
> 0.57
> Sol_RF 30 124.15 30.00 179.99 30.00 -111.71 150.00 1147.
> 0.57
>
> My question is:
> 1. For a 70 kDa protein compelx, is it common to have such a large cell
> with a dimention as long as 480 angstrom?
> 2. Is it possible that the longest dimention of cell is doubled? If it is,
> how to divide it?
> 3. How to interpret the self-rotation results. The results from CNS and
> Molrep differs so much.
> 4. Any other suggestions on the molecular replacement are appraciated.
>
> Thanks.
>
> Wei Zhang
> PKU
>
>
>
> Lijun Liu, PhD
> Institute of Molecular Biology
> Department of Physics
> University of Oregon
> Eugene, OR 97403
> 541-346-5176
> http://www.uoregon.edu/~liulj/ <http://www.uoregon.edu/%7Eliulj/>
>
>
>
>
>
>
>
>
