Re: [ccp4bb] refmac5.7 refine pesudo-translational symmetry

[Qixu Cai]

Dear Eleanor,

The cell dimensions of the dataset with tNCS are 63.995   75.459  128.860
90.00  90.00  90.00 in P 2 21 21 space group.

But for the other dataset without tNCS of the same protein, the cell
dimensions are 64.203   65.319   76.443  90.00  90.00  90.00 in C 2 2 21.

So the cell volume of C2221 is 25% of the P22121. But I cannot index the
data to C2221 space group.

Thanks a lot.

Best wishes,

Qixu Cai


2012/8/1 Eleanor Dodson 

> Are your unit cell and SG correct? I think you maybe should reindex to get
> a cell volume 25% of this one, and maybe SG P21
> That patterson peak is enormous..
> Eleanor
> On 1 Aug 2012, at 08:45, Qixu Cai wrote:
>
> Dear Randy,
>
> Thanks very much for your detailed explanation and helpful advice. I have
> run the phaser job just as you have said. This is the result.
>
> The resolution of this dataset is 2.45A.
>
> =
>
> I added the three commands to phaser job for all alternative space groupof 
> P212121:
>
> TNCS USE ON
> TNCS NMOL 4
> TNCS PATT PERCENT 80.0
>
> The phaser got a SINGLE solution for space group P22121, and Rval=88.2
>
>SOLU SET  RFZ=14.5 TFZ=22.3 PAK=0 +TNCS PAK=0 +TNCS PAK=0 +TNCS PAK=0
> LLG=2565LLG=4322
>SOLU SPAC P 2 21 21
>SOLU 6DIM ENSE ensemble1 EULER 91.7 89.4 90.3 FRAC 0.49 0.51 0.99 BFAC
> 0.00
>SOLU 6DIM ENSE ensemble1 EULER 269.3 89.4 90.2 FRAC -0.01 -0.00 0.74
> BFAC 0.00
>SOLU 6DIM ENSE ensemble1 EULER 269.0 89.1 90.0 FRAC 0.49 -0.00 1.00
> BFAC 0.00
>SOLU 6DIM ENSE ensemble1 EULER 91.5 89.3 90.2 FRAC 0.99 0.51 0.74 BFAC
> 0.00
>
> After 50 cycles rigidbody refinement, R/Rfree=0.36/0.37
> After 50 cycles restraint refinement (with jellybody refine and twin
> refine), R/Rfree=0.31/0.35
> After several cycles of coot model building and restraint refinement,
> R/Rfree=0.27/0.31
> After finding waters, R/Rfree=0.2478/0.2984
>
> ===
> If I run phaser job without those three added commands, the phaser got
> single solution at P21212 space group (Rval=0.3%):
>
>
>SOLU SET  RFZ=17.0 TFZ=20.1 PAK=0 LLG=457 TFZ==16.2 RFZ=16.3 TFZ=62.1
> PAK=0
> LLG=2599 TFZ==19.8 (& TFZ=59.0 PAK=0 LLG=2403) LLG+=(2608 & 4349)
> LLG=4200
> TFZ==22.2 PAK=0 RFZ=10.8 TFZ=56.4 PAK=0 LLG=5751 TFZ==22.5 LLG=9274
> TFZ==27.5
>SOLU SPAC P 21 21 2
>SOLU 6DIM ENSE ensemble1 EULER 90.0 90.7 270.1 FRAC -0.00 0.26 0.13
> BFAC -4.09
>SOLU 6DIM ENSE ensemble1 EULER 271.2 89.4 90.3 FRAC 0.01 0.24 -0.38
> BFAC -3.00
>SOLU 6DIM ENSE ensemble1 EULER 270.3 90.8 270.1 FRAC 0.50 -0.25 -0.12
> BFAC -1.25
>SOLU 6DIM ENSE ensemble1 EULER 87.5 90.6 270.4 FRAC 0.02 0.26 -0.37
> BFAC 11.58
>
> And the electron density is worse than the P22121 solution.
>
> ==
> Just as I have said last email, I can also get single solution at P212121
> space group,
>
> and after 50 cycles rigidbody refinement and 50 cycles restraint
> refinement with jellybody and twin, I can get R/Rfree=0.30/0.33.
>
> But the electron density is worse than the P22121 solution, so I do not
> carry out the model building in coot.
>
> ===
>
> My questiones:
>
> 1) Is the P22121 solution is my correct solution? These three solutions
> really confused me.
>
> 2) Why the R/Rfree is high even after I have good electron density and
> have found waters? (R/Rfree=0.478/0.2984 for 2.45A data)
>
> 3) What's the function of the command "TNCS USE ON"? Is it necessary?
>
> 4) I found if I used twin refinement in refmac5, the R/Rfree would
> decrease about 0.02 comparing to without twinn refinement. Is it
> reasonable? Is there any tNCS refinement options in refmac?
>
> Thanks for your help.
>
> Best wishes,
>
> Qixu Cai
>
>
>
>
> 2012/7/31 Randy Read 
>
>> Hi,
>>
>> The second Patterson peak is twice the first (considering lattice
>> translations, where 1 is equivalent to 0 modulo 1), and then if you triple
>> the first vector you'll get minus the first vector (again considering
>> lattice translations, i.e. 3/4 is equal to 1 - 1/4 which is equivalent to
>> -1/4), which is equivalent by symmetry to the first vector so wouldn't
>> appear in the peak list.  So the Patterson indicates 4 copies separated by
>> 0, 1, 2 and 3 times the top Patterson vector, in approximately the same
>> orientation.
>>
>> We've haven't fully dealt with the complications of multiple tNCS-related
>> copies in Phaser yet, but for this type of case there is a reasonable
>> treatment.  You should add two commands to the Phaser job:
>>
>> TNCS NMOL 4
>> TNCS PATT PERCENT 80
>>
>> The first says that the Patterson translation is repeated 4 times, and
>> the second will cause the second Patterson peak to be ignored.
>>
>> I'd suggest repeating the Phaser run with these commands and making sure
>> that you end up with the same solution as you got wh

Re: [ccp4bb] refmac5.7 refine pesudo-translational symmetry

[Eleanor Dodson]

Are your unit cell and SG correct? I think you maybe should reindex to get a 
cell volume 25% of this one, and maybe SG P21
That patterson peak is enormous..
Eleanor
On 1 Aug 2012, at 08:45, Qixu Cai wrote:

> Dear Randy,
> 
> Thanks very much for your detailed explanation and helpful advice. I have run 
> the phaser job just as you have said. This is the result.
> 
> The resolution of this dataset is 2.45A.
> 
> =
> 
> I added the three commands to phaser job for all alternative space group of 
> P212121:
> 
> TNCS USE ON
> TNCS NMOL 4
> TNCS PATT PERCENT 80.0
> 
> The phaser got a SINGLE solution for space group P22121, and Rval=88.2
> 
>SOLU SET  RFZ=14.5 TFZ=22.3 PAK=0 +TNCS PAK=0 +TNCS PAK=0 +TNCS PAK=0 
> LLG=2565LLG=4322
>SOLU SPAC P 2 21 21
>SOLU 6DIM ENSE ensemble1 EULER 91.7 89.4 90.3 FRAC 0.49 0.51 0.99 BFAC 0.00
>SOLU 6DIM ENSE ensemble1 EULER 269.3 89.4 90.2 FRAC -0.01 -0.00 0.74 BFAC 
> 0.00
>SOLU 6DIM ENSE ensemble1 EULER 269.0 89.1 90.0 FRAC 0.49 -0.00 1.00 BFAC 
> 0.00
>SOLU 6DIM ENSE ensemble1 EULER 91.5 89.3 90.2 FRAC 0.99 0.51 0.74 BFAC 0.00
> 
> After 50 cycles rigidbody refinement, R/Rfree=0.36/0.37
> After 50 cycles restraint refinement (with jellybody refine and twin refine), 
> R/Rfree=0.31/0.35
> After several cycles of coot model building and restraint refinement, 
> R/Rfree=0.27/0.31
> After finding waters, R/Rfree=0.2478/0.2984
> 
> ===
> If I run phaser job without those three added commands, the phaser got single 
> solution at P21212 space group (Rval=0.3%):
> 
> 
>SOLU SET  RFZ=17.0 TFZ=20.1 PAK=0 LLG=457 TFZ==16.2 RFZ=16.3 TFZ=62.1 PAK=0
> LLG=2599 TFZ==19.8 (& TFZ=59.0 PAK=0 LLG=2403) LLG+=(2608 & 4349) LLG=4200
> TFZ==22.2 PAK=0 RFZ=10.8 TFZ=56.4 PAK=0 LLG=5751 TFZ==22.5 LLG=9274 
> TFZ==27.5
>SOLU SPAC P 21 21 2
>SOLU 6DIM ENSE ensemble1 EULER 90.0 90.7 270.1 FRAC -0.00 0.26 0.13 BFAC 
> -4.09
>SOLU 6DIM ENSE ensemble1 EULER 271.2 89.4 90.3 FRAC 0.01 0.24 -0.38 BFAC 
> -3.00
>SOLU 6DIM ENSE ensemble1 EULER 270.3 90.8 270.1 FRAC 0.50 -0.25 -0.12 BFAC 
> -1.25
>SOLU 6DIM ENSE ensemble1 EULER 87.5 90.6 270.4 FRAC 0.02 0.26 -0.37 BFAC 
> 11.58
> 
> And the electron density is worse than the P22121 solution.
> 
> ==
> Just as I have said last email, I can also get single solution at P212121 
> space group,
> 
> and after 50 cycles rigidbody refinement and 50 cycles restraint refinement 
> with jellybody and twin, I can get R/Rfree=0.30/0.33.
> 
> But the electron density is worse than the P22121 solution, so I do not carry 
> out the model building in coot.
> 
> ===
> 
> My questiones:
> 
> 1) Is the P22121 solution is my correct solution? These three solutions 
> really confused me.
> 
> 2) Why the R/Rfree is high even after I have good electron density and have 
> found waters? (R/Rfree=0.478/0.2984 for 2.45A data)
> 
> 3) What's the function of the command "TNCS USE ON"? Is it necessary?
> 
> 4) I found if I used twin refinement in refmac5, the R/Rfree would decrease 
> about 0.02 comparing to without twinn refinement. Is it reasonable? Is there 
> any tNCS refinement options in refmac?
> 
> Thanks for your help.
> 
> Best wishes,
> 
> Qixu Cai
> 
> 
> 
> 
> 2012/7/31 Randy Read 
> Hi,
> 
> The second Patterson peak is twice the first (considering lattice 
> translations, where 1 is equivalent to 0 modulo 1), and then if you triple 
> the first vector you'll get minus the first vector (again considering lattice 
> translations, i.e. 3/4 is equal to 1 - 1/4 which is equivalent to
> -1/4), which is equivalent by symmetry to the first vector so wouldn't appear 
> in the peak list.  So the Patterson indicates 4 copies separated by 0, 1, 2 
> and 3 times the top Patterson vector, in approximately the same orientation.
> 
> We've haven't fully dealt with the complications of multiple tNCS-related 
> copies in Phaser yet, but for this type of case there is a reasonable 
> treatment.  You should add two commands to the Phaser job:
> 
> TNCS NMOL 4
> TNCS PATT PERCENT 80
> 
> The first says that the Patterson translation is repeated 4 times, and the 
> second will cause the second Patterson peak to be ignored.
> 
> I'd suggest repeating the Phaser run with these commands and making sure that 
> you end up with the same solution as you got when the tNCS was ignored.  When 
> tNCS is ignored, it's possible to end up with a solution that is only 
> partially correct, which would be one explanation for having some molecules 
> that look better in density than others.
> 
> Best wishes,
> 
> Randy Read
> 
> On 31 Jul 2012, at 13:11, Qixu Cai wrote:
> 
> > It's a P212121 dataset. I have used phaser to find four solution in ASU.
> >
> > This is the phaser log file:
> > -

Re: [ccp4bb] refmac5.7 refine pesudo-translational symmetry

[Qixu Cai]

Dear Randy,

Thanks very much for your detailed explanation and helpful advice. I have
run the phaser job just as you have said. This is the result.

The resolution of this dataset is 2.45A.

=

I added the three commands to phaser job for all alternative space group of
P212121:

TNCS USE ON
TNCS NMOL 4
TNCS PATT PERCENT 80.0

The phaser got a SINGLE solution for space group P22121, and Rval=88.2

   SOLU SET  RFZ=14.5 TFZ=22.3 PAK=0 +TNCS PAK=0 +TNCS PAK=0 +TNCS PAK=0
LLG=2565LLG=4322
   SOLU SPAC P 2 21 21
   SOLU 6DIM ENSE ensemble1 EULER 91.7 89.4 90.3 FRAC 0.49 0.51 0.99 BFAC
0.00
   SOLU 6DIM ENSE ensemble1 EULER 269.3 89.4 90.2 FRAC -0.01 -0.00 0.74
BFAC 0.00
   SOLU 6DIM ENSE ensemble1 EULER 269.0 89.1 90.0 FRAC 0.49 -0.00 1.00 BFAC
0.00
   SOLU 6DIM ENSE ensemble1 EULER 91.5 89.3 90.2 FRAC 0.99 0.51 0.74 BFAC
0.00

After 50 cycles rigidbody refinement, R/Rfree=0.36/0.37
After 50 cycles restraint refinement (with jellybody refine and twin
refine), R/Rfree=0.31/0.35
After several cycles of coot model building and restraint refinement,
R/Rfree=0.27/0.31
After finding waters, R/Rfree=0.2478/0.2984

===
If I run phaser job without those three added commands, the phaser got
single solution at P21212 space group (Rval=0.3%):


   SOLU SET  RFZ=17.0 TFZ=20.1 PAK=0 LLG=457 TFZ==16.2 RFZ=16.3 TFZ=62.1
PAK=0
LLG=2599 TFZ==19.8 (& TFZ=59.0 PAK=0 LLG=2403) LLG+=(2608 & 4349)
LLG=4200
TFZ==22.2 PAK=0 RFZ=10.8 TFZ=56.4 PAK=0 LLG=5751 TFZ==22.5 LLG=9274
TFZ==27.5
   SOLU SPAC P 21 21 2
   SOLU 6DIM ENSE ensemble1 EULER 90.0 90.7 270.1 FRAC -0.00 0.26 0.13 BFAC
-4.09
   SOLU 6DIM ENSE ensemble1 EULER 271.2 89.4 90.3 FRAC 0.01 0.24 -0.38 BFAC
-3.00
   SOLU 6DIM ENSE ensemble1 EULER 270.3 90.8 270.1 FRAC 0.50 -0.25 -0.12
BFAC -1.25
   SOLU 6DIM ENSE ensemble1 EULER 87.5 90.6 270.4 FRAC 0.02 0.26 -0.37 BFAC
11.58

And the electron density is worse than the P22121 solution.

==
Just as I have said last email, I can also get single solution at P212121
space group,

and after 50 cycles rigidbody refinement and 50 cycles restraint refinement
with jellybody and twin, I can get R/Rfree=0.30/0.33.

But the electron density is worse than the P22121 solution, so I do not
carry out the model building in coot.

===

My questiones:

1) Is the P22121 solution is my correct solution? These three solutions
really confused me.

2) Why the R/Rfree is high even after I have good electron density and have
found waters? (R/Rfree=0.478/0.2984 for 2.45A data)

3) What's the function of the command "TNCS USE ON"? Is it necessary?

4) I found if I used twin refinement in refmac5, the R/Rfree would decrease
about 0.02 comparing to without twinn refinement. Is it reasonable? Is
there any tNCS refinement options in refmac?

Thanks for your help.

Best wishes,

Qixu Cai




2012/7/31 Randy Read 

> Hi,
>
> The second Patterson peak is twice the first (considering lattice
> translations, where 1 is equivalent to 0 modulo 1), and then if you triple
> the first vector you'll get minus the first vector (again considering
> lattice translations, i.e. 3/4 is equal to 1 - 1/4 which is equivalent to
> -1/4), which is equivalent by symmetry to the first vector so wouldn't
> appear in the peak list.  So the Patterson indicates 4 copies separated by
> 0, 1, 2 and 3 times the top Patterson vector, in approximately the same
> orientation.
>
> We've haven't fully dealt with the complications of multiple tNCS-related
> copies in Phaser yet, but for this type of case there is a reasonable
> treatment.  You should add two commands to the Phaser job:
>
> TNCS NMOL 4
> TNCS PATT PERCENT 80
>
> The first says that the Patterson translation is repeated 4 times, and the
> second will cause the second Patterson peak to be ignored.
>
> I'd suggest repeating the Phaser run with these commands and making sure
> that you end up with the same solution as you got when the tNCS was
> ignored.  When tNCS is ignored, it's possible to end up with a solution
> that is only partially correct, which would be one explanation for having
> some molecules that look better in density than others.
>
> Best wishes,
>
> Randy Read
>
> On 31 Jul 2012, at 13:11, Qixu Cai wrote:
>
> > It's a P212121 dataset. I have used phaser to find four solution in ASU.
> >
> > This is the phaser log file:
> > 
> > PEUDO-TRANSLATIONAL NCS VECTOR
> > --
> >
> >   Space Group :   P 21 21 21
> >   Patterson Symmetry: P m m m
> >   Resolution of All Data (Number):2.45  49.00 (22968)
> >   Resolution of Patterson (Number):   5.00   9.99 (2364)
> >   There were 2 non-origin distinct peaks (i.e. more than 1

Re: [ccp4bb] refmac5.7 refine pesudo-translational symmetry

[Randy Read]

Hi,

The second Patterson peak is twice the first (considering lattice translations, 
where 1 is equivalent to 0 modulo 1), and then if you triple the first vector 
you'll get minus the first vector (again considering lattice translations, i.e. 
3/4 is equal to 1 - 1/4 which is equivalent to 
-1/4), which is equivalent by symmetry to the first vector so wouldn't appear 
in the peak list.  So the Patterson indicates 4 copies separated by 0, 1, 2 and 
3 times the top Patterson vector, in approximately the same orientation.

We've haven't fully dealt with the complications of multiple tNCS-related 
copies in Phaser yet, but for this type of case there is a reasonable 
treatment.  You should add two commands to the Phaser job:

TNCS NMOL 4
TNCS PATT PERCENT 80

The first says that the Patterson translation is repeated 4 times, and the 
second will cause the second Patterson peak to be ignored.

I'd suggest repeating the Phaser run with these commands and making sure that 
you end up with the same solution as you got when the tNCS was ignored.  When 
tNCS is ignored, it's possible to end up with a solution that is only partially 
correct, which would be one explanation for having some molecules that look 
better in density than others.

Best wishes,

Randy Read

On 31 Jul 2012, at 13:11, Qixu Cai wrote:

> It's a P212121 dataset. I have used phaser to find four solution in ASU.
> 
> This is the phaser log file:
> 
> PEUDO-TRANSLATIONAL NCS VECTOR
> --
> 
>   Space Group :   P 21 21 21
>   Patterson Symmetry: P m m m
>   Resolution of All Data (Number):2.45  49.00 (22968)
>   Resolution of Patterson (Number):   5.00   9.99 (2364)
>   There were 2 non-origin distinct peaks (i.e. more than 15 angstroms from the
>   origin)
> 
> 
>   84.1% origin:   FRAC 0.500 0.000 0.250   (ORTH   32.00.0   32.2)
>   72.2% origin:   FRAC 0.000 0.000 0.500   (ORTH0.00.0   64.4)
> 
> 
>   More than one pseudo-translational ncs vector found
>  Correction factors will not be applied
> 
> 
> PS: I have used phenix.xtrige and found the p-value of
> pseudo-translational ncs is very little, which indicates the exist of
> the pseudo-translational ncs. And no twin found in this dataset.
> 
> Now the problem is two in the four molecules of an ASU have worse
> electron density than the other two molecules. And after rigidbody and
> restraint refinement by refmac without "twin refinement", the R/Rfree
> is a little high (0.33/0.36). And if I turn on the "twin refinement"
> in refmac, the R/Rfree is 0.30/0.33.
> 
> So, my question is, there is not twin in my data but
> pseudo-translational ncs, is it suitable to use "twin refinement" in
> refmac, which has a good R/Rfree result.
> 
> Thanks a lot for your help.
> 
> Best wishes,
> 
> Qixu Cai
> 
> 
> 2012/7/31, Eleanor Dodson :
>> More details - what do you mean by pesudo-translational symmetry ?
>> Are there two molecules related by a translation vector? or its it
>> something more complicated?
>> Eleanor
>> 
>> On 31 July 2012 10:47, Qixu Cai  wrote:
>> 
>>> Dear all,
>>> 
>>> Can I use the "twin refinement" to refine the pesudo-translational
>>> symmetry dataset?
>>> 
>>> Thanks a lot for your help.
>>> 
>>> Best wishes,
>>> 
>>> Qixu Cai
>>> 
>> 
> 
> 
> -- 
> Qixu Cai
> Email: caiq...@gmail.com
> School of Life Sciences,
> Xiamen University, Fujian, China

--
Randy J. Read
Department of Haematology, University of Cambridge
Cambridge Institute for Medical Research  Tel: + 44 1223 336500
Wellcome Trust/MRC Building   Fax: + 44 1223 336827
Hills RoadE-mail: rj...@cam.ac.uk
Cambridge CB2 0XY, U.K.   www-structmed.cimr.cam.ac.uk

Re: [ccp4bb] refmac5.7 refine pesudo-translational symmetry

[Qixu Cai]

It's a P212121 dataset. I have used phaser to find four solution in ASU.

This is the phaser log file:

PEUDO-TRANSLATIONAL NCS VECTOR
--

   Space Group :   P 21 21 21
   Patterson Symmetry: P m m m
   Resolution of All Data (Number):2.45  49.00 (22968)
   Resolution of Patterson (Number):   5.00   9.99 (2364)
   There were 2 non-origin distinct peaks (i.e. more than 15 angstroms from the
   origin)


   84.1% origin:   FRAC 0.500 0.000 0.250   (ORTH   32.00.0   32.2)
   72.2% origin:   FRAC 0.000 0.000 0.500   (ORTH0.00.0   64.4)


   More than one pseudo-translational ncs vector found
  Correction factors will not be applied


PS: I have used phenix.xtrige and found the p-value of
pseudo-translational ncs is very little, which indicates the exist of
the pseudo-translational ncs. And no twin found in this dataset.

Now the problem is two in the four molecules of an ASU have worse
electron density than the other two molecules. And after rigidbody and
restraint refinement by refmac without "twin refinement", the R/Rfree
is a little high (0.33/0.36). And if I turn on the "twin refinement"
in refmac, the R/Rfree is 0.30/0.33.

So, my question is, there is not twin in my data but
pseudo-translational ncs, is it suitable to use "twin refinement" in
refmac, which has a good R/Rfree result.

Thanks a lot for your help.

Best wishes,

Qixu Cai


2012/7/31, Eleanor Dodson :
> More details - what do you mean by pesudo-translational symmetry ?
> Are there two molecules related by a translation vector? or its it
> something more complicated?
> Eleanor
>
> On 31 July 2012 10:47, Qixu Cai  wrote:
>
>> Dear all,
>>
>> Can I use the "twin refinement" to refine the pesudo-translational
>> symmetry dataset?
>>
>> Thanks a lot for your help.
>>
>> Best wishes,
>>
>> Qixu Cai
>>
>


-- 
Qixu Cai
Email: caiq...@gmail.com
School of Life Sciences,
Xiamen University, Fujian, China

Re: [ccp4bb] refmac5.7 refine pesudo-translational symmetry

[Steiner, Roberto]

My understanding is that ML functions in the presence of pseudo-translational 
symmetry are suboptimal (see a very short discussion in Acta Cryst. (2011). 
D67, 355–367. Acta Cryst. (2008). D64, 99–107 is also a good reading).
Having said that if your crystal/dataset exhibits twinning on top of 
pseduo-translation you should activate the twin refinement option in Refmac.

Roberto

On 31 Jul 2012, at 10:47, Qixu Cai wrote:

Dear all,

Can I use the "twin refinement" to refine the pesudo-translational symmetry 
dataset?

Thanks a lot for your help.

Best wishes,

Qixu Cai

Roberto Steiner
Randall Division of Cell and Molecular Biophysics
King's College London

Room 3.10A
New Hunt's House
Guy's Campus
SE1 1UL, London, UK
Tel 0044-20-78488216
Fax 0044-20-78486435
roberto.stei...@kcl.ac.uk

[ccp4bb] refmac5.7 refine pesudo-translational symmetry

[Qixu Cai]

Dear all,

Can I use the "twin refinement" to refine the pesudo-translational symmetry
dataset?

Thanks a lot for your help.

Best wishes,

Qixu Cai