Thanks a lot Elaine. I will try to work on it now and see if I can get any results.

Regards, Haresh

On Mon, Jan 27, 2014 at 12:28 PM, Elaine Meng meng@cgl.ucsf.edu wrote:

Hi Haresh, The Reply Log message (see the bottom part of what you sent) says there was a problem calculating the charges of your mutated residues and that you should try running Add Charge first, so that's what I would recommend. Add Charge is in the menu under Tools.. Structure Editing, or can be run with command "addcharge". It has two options for calculating the charges of nonstandard residues: AM1-BCC or Gasteiger. Since the message shows there was a problem calculating using the AM1-BCC method on your mutated residues, I would try the Gasteiger method instead.

< http://www.rbvi.ucsf.edu/chimera/docs/ContributedSoftware/addcharge/addcharg...

...

http://www.rbvi.ucsf.edu/chimera/docs/UsersGuide/midas/addcharge.html

Then, after you run Add Charge successfully, you can run Coulombic again. It will automatically detect that the charges have already been calculated, and use those charges for calculating the ESP and coloring the surface.

Actually I tried calculating charges for a single 5,5,5-trifluoroleucine residue by itself (in Chimera, can use "open pubchem:259832" to fetch it by PubChem CID) <http://pubchem.ncbi.nlm.nih.gov/summary/summary.cgi?cid=259832&loc=ec_rc...

...

...and both methods, AM1-BCC and Gasteiger, worked fine; however maybe the calculation is more difficult in your structure where it sounds like the two mutated residues are treated together.

I hope this helps, Elaine

---

Elaine C. Meng, Ph.D. UCSF Computer Graphics Lab (Chimera team) and Babbitt Lab Department of Pharmaceutical Chemistry University of California, San Francisco

On Jan 26, 2014, at 8:52 PM, Haresh Tukaram More htm211@nyu.edu wrote:

...

Hi, I tried to get the Coulombic surface coloring for one of the protein

that I modified by removing certain residue at the C-termini and then mutated some residues to the non-natural amino acid 5,5,5-trifluoroleucine using Swisssidechin option. I am getting this error which I am not able to understand.

...

Disabled GPU programs because a graphics driver bug was encountered while compiling a vertex shader.

#0, chain A: COMP #0, chain B: COMP #0, chain C: COMP #0, chain D: COMP #0, chain E: COMP C:\Program Files\Chimera 1.8.1\bin\mscalc.exe 1.400000 2.000000 1 MSMSLIB 1.3 started on Local PC Copyright M.F. Sanner (March 2000) Compilation flags

Surface CC TFLE.pdb, category main, probe radius 1.4, vertex density 2 22 connected surface components Total solvent excluded surface area = 10393.6 component areas = 8599.46, 624.842, 225.9, 216.153, 81.7059,

54.8894, 54.839, 48.0624, 45.9525, 43.9588, 41.5667, 39.1204, 38.7549, 37.0513, 35.5418, 33.6712, 32.9167, 28.9293, 27.8262, 27.7523, 27.7319, 26.9429

...

Total solvent accessible surface area = 10372 component areas = 9971.67, 202.012, 92.2225, 70.7414, 13.8165,

4.12717, 3.62753, 3.24856, 2.597, 1.1866, 1.72654, 1.33137, 1.08737, 0.962375, 0.64812, 0.231422, 0.40193, 0.120068, 0.0570046, 0.0515394, 0.0674059, 0.0311705

...

No SEQRES records for combination (#-2147483648.-2147483648) chain A;

guessing terminii instead

...

No SEQRES records for combination (#-2147483648.-2147483648) chain B;

guessing terminii instead

...

No SEQRES records for combination (#-2147483648.-2147483648) chain C;

guessing terminii instead

...

No SEQRES records for combination (#-2147483648.-2147483648) chain D;

guessing terminii instead

...

No SEQRES records for combination (#-2147483648.-2147483648) chain E;

guessing terminii instead

...

Chain-initial residues that are actual N terminii:

#-2147483648.-2147483648 MET 27.A, #-2147483648.-2147483648 MET 27.B, #-2147483648.-2147483648 MET 27.C, #-2147483648.-2147483648 MET 27.D, #-2147483648.-2147483648 MET 27.E

...

Chain-initial residues that are not actual N terminii: Chain-final residues that are actual C terminii:

#-2147483648.-2147483648 TFLE 67.A, #-2147483648.-2147483648 TFLE 67.B, #-2147483648.-2147483648 TFLE 67.C, #-2147483648.-2147483648 TFLE 67.D, #-2147483648.-2147483648 TFLE 67.E

...

Chain-final residues that are not actual C terminii: 424 hydrogen bonds Hydrogens added Charge model: AMBER ff12SB Assigning partial charges to residue TFLE+TFLE (net charge +0) with

am1-bcc method

...

Running ANTECHAMBER command: C:/Program Files/Chimera

1.8.1/bin/amber12/bin\antechamber -ek qm_theory='AM1', -i c:\users\haresh~1.pol\appdata\local\temp\tmpipssqi\ante.in.mol2 -fi mol2 -o c:\users\haresh~1.pol\appdata\local\temp\tmpipssqi\ante.out.mol2 -fo mol2 -c bcc -nc 0 -j 5 -s 2

...

(TFLE+TFLE)

Cannot automatically determine charges for residue TFLE+TFLE; Run AddCharge tool manually to add charges and then rerun ESP

Can someone please tell me a step by step procedure to make this work. Thanks & Regards, Haresh

HI Elaine,

I tried to do that, but not able to add charges manually. I have attached a PDB filer here with TFLE mutations. Can you please see if you can add charges on it. I am unable to give charge to TFLE as all the time the process stop at antechamber.exe and abruptly closes the chimera.

Thanks & Regards, Haresh

On Tue, Jan 28, 2014 at 12:27 PM, Elaine Meng meng@cgl.ucsf.edu wrote:

Hi Haresh, There is another issue you may wish to consider: Add Charge will require adding hydrogens first, but then the molecular surface will be more "craggy" due to the additional small atoms. Here are instructions if you want to show the Coulombic ESP coloring on the less rugged surface calculated without hydrogens (as is usually done; radii automatically adjust to compensate):

After adding hydrogens and charges: (1) when you use the Coulombic Surface Coloring tool, turn on the option to "Compute grid..." (in my test just now I took the defaults for the other grid options)

(2) delete the hydrogens, e.g. command: del H

(3) use the "Surface Color" dialog that automatically appeared when you created the grid to color the main surface by electrostatic potential using the potential file (grid) you just created. If you don't see that dialog, you can raise it using menu: Tools... Surface/Binding Analysis... Electrostatic Surface Coloring. The default name of that grid is Coulombic ESP. (Do not use the Coulombic Surface Coloring dialog for coloring! After the hydrogens with many positive charges have been deleted, mostly negative charges will be left, and you will get an incorrect very red surface using the Coulombic Surface Coloring dialog.)

Normally the protein wouldn't have strange residues that require a separate charge calculation, and this circuitous process wouldn't be required. Best, Elaine

---

Elaine C. Meng, Ph.D. UCSF Computer Graphics Lab (Chimera team) and Babbitt Lab Department of Pharmaceutical Chemistry University of California, San Francisco

On Jan 27, 2014, at 6:56 PM, Haresh Tukaram More htm211@nyu.edu wrote:

...

Thanks a lot Elaine. I will try to work on it now and see if I can get

any results.

...

Regards, Haresh

On Mon, Jan 27, 2014 at 12:28 PM, Elaine Meng meng@cgl.ucsf.edu wrote: Hi Haresh, The Reply Log message (see the bottom part of what you sent) says there

was a problem calculating the charges of your mutated residues and that you should try running Add Charge first, so that's what I would recommend. Add Charge is in the menu under Tools.. Structure Editing, or can be run with command "addcharge". It has two options for calculating the charges of nonstandard residues: AM1-BCC or Gasteiger. Since the message shows there was a problem calculating using the AM1-BCC method on your mutated residues, I would try the Gasteiger method instead.

...

<

http://www.rbvi.ucsf.edu/chimera/docs/ContributedSoftware/addcharge/addcharg...

...

http://www.rbvi.ucsf.edu/chimera/docs/UsersGuide/midas/addcharge.html

Then, after you run Add Charge successfully, you can run Coulombic

again. It will automatically detect that the charges have already been calculated, and use those charges for calculating the ESP and coloring the surface.

...

Actually I tried calculating charges for a single 5,5,5-trifluoroleucine

residue by itself (in Chimera, can use "open pubchem:259832" to fetch it by PubChem CID)

...

<

http://pubchem.ncbi.nlm.nih.gov/summary/summary.cgi?cid=259832&loc=ec_rc...

...

...and both methods, AM1-BCC and Gasteiger, worked fine; however maybe

the calculation is more difficult in your structure where it sounds like the two mutated residues are treated together.

...

I hope this helps, Elaine

---

Elaine C. Meng, Ph.D. UCSF Computer Graphics Lab (Chimera team) and Babbitt Lab Department of Pharmaceutical Chemistry University of California, San Francisco

On Jan 26, 2014, at 8:52 PM, Haresh Tukaram More htm211@nyu.edu wrote:

...

Hi, I tried to get the Coulombic surface coloring for one of the protein

that I modified by removing certain residue at the C-termini and then mutated some residues to the non-natural amino acid 5,5,5-trifluoroleucine using Swisssidechin option. I am getting this error which I am not able to understand.

...

...

Disabled GPU programs because a graphics driver bug was encountered while compiling a vertex shader.

#0, chain A: COMP #0, chain B: COMP #0, chain C: COMP #0, chain D: COMP #0, chain E: COMP C:\Program Files\Chimera 1.8.1\bin\mscalc.exe 1.400000 2.000000 1 MSMSLIB 1.3 started on Local PC Copyright M.F. Sanner (March 2000) Compilation flags

Surface CC TFLE.pdb, category main, probe radius 1.4, vertex density 2 22 connected surface components Total solvent excluded surface area = 10393.6 component areas = 8599.46, 624.842, 225.9, 216.153, 81.7059,

54.8894, 54.839, 48.0624, 45.9525, 43.9588, 41.5667, 39.1204, 38.7549, 37.0513, 35.5418, 33.6712, 32.9167, 28.9293, 27.8262, 27.7523, 27.7319, 26.9429

...

...

Total solvent accessible surface area = 10372 component areas = 9971.67, 202.012, 92.2225, 70.7414, 13.8165,

4.12717, 3.62753, 3.24856, 2.597, 1.1866, 1.72654, 1.33137, 1.08737, 0.962375, 0.64812, 0.231422, 0.40193, 0.120068, 0.0570046, 0.0515394, 0.0674059, 0.0311705

...

...

No SEQRES records for combination (#-2147483648.-2147483648) chain A;

guessing terminii instead

...

...

No SEQRES records for combination (#-2147483648.-2147483648) chain B;

guessing terminii instead

...

...

No SEQRES records for combination (#-2147483648.-2147483648) chain C;

guessing terminii instead

...

...

No SEQRES records for combination (#-2147483648.-2147483648) chain D;

guessing terminii instead

...

...

No SEQRES records for combination (#-2147483648.-2147483648) chain E;

guessing terminii instead

...

...

Chain-initial residues that are actual N terminii:

#-2147483648.-2147483648 MET 27.A, #-2147483648.-2147483648 MET 27.B, #-2147483648.-2147483648 MET 27.C, #-2147483648.-2147483648 MET 27.D, #-2147483648.-2147483648 MET 27.E

...

...

Chain-initial residues that are not actual N terminii: Chain-final residues that are actual C terminii:

#-2147483648.-2147483648 TFLE 67.A, #-2147483648.-2147483648 TFLE 67.B, #-2147483648.-2147483648 TFLE 67.C, #-2147483648.-2147483648 TFLE 67.D, #-2147483648.-2147483648 TFLE 67.E

...

...

Chain-final residues that are not actual C terminii: 424 hydrogen bonds Hydrogens added Charge model: AMBER ff12SB Assigning partial charges to residue TFLE+TFLE (net charge +0) with

am1-bcc method

...

...

Running ANTECHAMBER command: C:/Program Files/Chimera

1.8.1/bin/amber12/bin\antechamber -ek qm_theory='AM1', -i c:\users\haresh~1.pol\appdata\local\temp\tmpipssqi\ante.in.mol2 -fi mol2 -o c:\users\haresh~1.pol\appdata\local\temp\tmpipssqi\ante.out.mol2 -fo mol2 -c bcc -nc 0 -j 5 -s 2

...

...

(TFLE+TFLE)

Cannot automatically determine charges for residue TFLE+TFLE; Run AddCharge tool manually to add charges and then rerun ESP

Can someone please tell me a step by step procedure to make this work. Thanks & Regards, Haresh

---

Chimera-users mailing list Chimera-users@cgl.ucsf.edu http://plato.cgl.ucsf.edu/mailman/listinfo/chimera-users

Hi Elaine,

Finally!! it worked. In the reply log it shows following

Charges for residue TFLE determined Assigning partial charges to residue CL (net charge -1) with gasteiger method Total charge for #0: 9.000

So i should use +9 charge for TFLE and TFLE+TFLE? I did that and now able to get the electrostatic surface potential. The way I did is as follows

1. AddH 2. Add charge 3. Select surface of protein 4. Select coulombic surface coloring command and selected the grid option. 5. Next the surface color box opened and I selected color option. 6. Once it is done I removed hydrogen by del H

and I got the protein with electrostatic surface potential.

I hope now this is correct.

Regards, Haresh

On Tue, Jan 28, 2014 at 1:44 PM, Elaine Meng meng@cgl.ucsf.edu wrote:

Hi Haresh, If you use "Gasteiger" instead of "AM1-BCC" in the Add Charge dialog as suggested in my first reply, it works (at least for me, but I'm pretty sure it will for you too)! Elaine

On Jan 28, 2014, at 10:29 AM, Haresh Tukaram More htm211@nyu.edu wrote:

...

HI Elaine,

I tried to do that, but not able to add charges manually. I have

attached a PDB filer here with TFLE mutations. Can you please see if you can add charges on it. I am unable to give charge to TFLE as all the time the process stop at antechamber.exe and abruptly closes the chimera.

...

Thanks & Regards, Haresh

On Tue, Jan 28, 2014 at 12:27 PM, Elaine Meng meng@cgl.ucsf.edu wrote: Hi Haresh, There is another issue you may wish to consider: Add Charge will

require adding hydrogens first, but then the molecular surface will be more "craggy" due to the additional small atoms. Here are instructions if you want to show the Coulombic ESP coloring on the less rugged surface calculated without hydrogens (as is usually done; radii automatically adjust to compensate):

...

After adding hydrogens and charges: (1) when you use the Coulombic Surface Coloring tool, turn on the option

to "Compute grid..." (in my test just now I took the defaults for the other grid options)

...

(2) delete the hydrogens, e.g. command: del H

(3) use the "Surface Color" dialog that automatically appeared when you

created the grid to color the main surface by electrostatic potential using the potential file (grid) you just created. If you don't see that dialog, you can raise it using menu: Tools... Surface/Binding Analysis... Electrostatic Surface Coloring. The default name of that grid is Coulombic ESP. (Do not use the Coulombic Surface Coloring dialog for coloring! After the hydrogens with many positive charges have been deleted, mostly negative charges will be left, and you will get an incorrect very red surface using the Coulombic Surface Coloring dialog.)

...

Normally the protein wouldn't have strange residues that require a

separate charge calculation, and this circuitous process wouldn't be required.

...

Best, Elaine

---

Elaine C. Meng, Ph.D. UCSF Computer Graphics Lab (Chimera team) and Babbitt Lab Department of Pharmaceutical Chemistry University of California, San Francisco

On Jan 27, 2014, at 6:56 PM, Haresh Tukaram More htm211@nyu.edu wrote:

...

Thanks a lot Elaine. I will try to work on it now and see if I can get

any results.

...

...

Regards, Haresh

On Mon, Jan 27, 2014 at 12:28 PM, Elaine Meng meng@cgl.ucsf.edu

wrote:

...

...

Hi Haresh, The Reply Log message (see the bottom part of what you sent) says

there was a problem calculating the charges of your mutated residues and that you should try running Add Charge first, so that's what I would recommend. Add Charge is in the menu under Tools.. Structure Editing, or can be run with command "addcharge". It has two options for calculating the charges of nonstandard residues: AM1-BCC or Gasteiger. Since the message shows there was a problem calculating using the AM1-BCC method on your mutated residues, I would try the Gasteiger method instead.

...

...

<

http://www.rbvi.ucsf.edu/chimera/docs/ContributedSoftware/addcharge/addcharg...

...

...

<http://www.rbvi.ucsf.edu/chimera/docs/UsersGuide/midas/addcharge.html

...

Then, after you run Add Charge successfully, you can run Coulombic

again. It will automatically detect that the charges have already been calculated, and use those charges for calculating the ESP and coloring the surface.

...

...

Actually I tried calculating charges for a single

5,5,5-trifluoroleucine residue by itself (in Chimera, can use "open pubchem:259832" to fetch it by PubChem CID)

...

...

<

http://pubchem.ncbi.nlm.nih.gov/summary/summary.cgi?cid=259832&loc=ec_rc...

...

...

...and both methods, AM1-BCC and Gasteiger, worked fine; however maybe

the calculation is more difficult in your structure where it sounds like the two mutated residues are treated together.

...

...

I hope this helps, Elaine

---

Elaine C. Meng, Ph.D. UCSF Computer Graphics Lab (Chimera team) and Babbitt Lab Department of Pharmaceutical Chemistry University of California, San Francisco

On Jan 26, 2014, at 8:52 PM, Haresh Tukaram More htm211@nyu.edu

wrote:

...

...

...

Hi, I tried to get the Coulombic surface coloring for one of the protein

that I modified by removing certain residue at the C-termini and then mutated some residues to the non-natural amino acid 5,5,5-trifluoroleucine using Swisssidechin option. I am getting this error which I am not able to understand.

...

...

...

Disabled GPU programs because a graphics driver bug was encountered while compiling a vertex shader.

#0, chain A: COMP #0, chain B: COMP #0, chain C: COMP #0, chain D: COMP #0, chain E: COMP C:\Program Files\Chimera 1.8.1\bin\mscalc.exe 1.400000 2.000000 1 MSMSLIB 1.3 started on Local PC Copyright M.F. Sanner (March 2000) Compilation flags

Surface CC TFLE.pdb, category main, probe radius 1.4, vertex density

2

...

...

...

22 connected surface components Total solvent excluded surface area = 10393.6 component areas = 8599.46, 624.842, 225.9, 216.153, 81.7059,

54.8894, 54.839, 48.0624, 45.9525, 43.9588, 41.5667, 39.1204, 38.7549, 37.0513, 35.5418, 33.6712, 32.9167, 28.9293, 27.8262, 27.7523, 27.7319, 26.9429

...

...

...

Total solvent accessible surface area = 10372 component areas = 9971.67, 202.012, 92.2225, 70.7414, 13.8165,

4.12717, 3.62753, 3.24856, 2.597, 1.1866, 1.72654, 1.33137, 1.08737, 0.962375, 0.64812, 0.231422, 0.40193, 0.120068, 0.0570046, 0.0515394, 0.0674059, 0.0311705

...

...

...

No SEQRES records for combination (#-2147483648.-2147483648) chain

A; guessing terminii instead

...

...

...

No SEQRES records for combination (#-2147483648.-2147483648) chain

B; guessing terminii instead

...

...

...

No SEQRES records for combination (#-2147483648.-2147483648) chain

C; guessing terminii instead

...

...

...

No SEQRES records for combination (#-2147483648.-2147483648) chain

D; guessing terminii instead

...

...

...

No SEQRES records for combination (#-2147483648.-2147483648) chain

E; guessing terminii instead

...

...

...

Chain-initial residues that are actual N terminii:

#-2147483648.-2147483648 MET 27.A, #-2147483648.-2147483648 MET 27.B, #-2147483648.-2147483648 MET 27.C, #-2147483648.-2147483648 MET 27.D, #-2147483648.-2147483648 MET 27.E

...

...

...

Chain-initial residues that are not actual N terminii: Chain-final residues that are actual C terminii:

#-2147483648.-2147483648 TFLE 67.A, #-2147483648.-2147483648 TFLE 67.B, #-2147483648.-2147483648 TFLE 67.C, #-2147483648.-2147483648 TFLE 67.D, #-2147483648.-2147483648 TFLE 67.E

...

...

...

Chain-final residues that are not actual C terminii: 424 hydrogen bonds Hydrogens added Charge model: AMBER ff12SB Assigning partial charges to residue TFLE+TFLE (net charge +0) with

am1-bcc method

...

...

...

Running ANTECHAMBER command: C:/Program Files/Chimera

1.8.1/bin/amber12/bin\antechamber -ek qm_theory='AM1', -i c:\users\haresh~1.pol\appdata\local\temp\tmpipssqi\ante.in.mol2 -fi mol2 -o c:\users\haresh~1.pol\appdata\local\temp\tmpipssqi\ante.out.mol2 -fo mol2 -c bcc -nc 0 -j 5 -s 2

...

...

...

(TFLE+TFLE)

Cannot automatically determine charges for residue TFLE+TFLE; Run AddCharge tool manually to add charges and then rerun ESP

Can someone please tell me a step by step procedure to make this

work.

...

...

...

Thanks & Regards, Haresh

Hi Elaine,

Yes I did exactly the way you explained and it is working now.

I have another question. I need to add some 14 residues on N-termius of the protein which I did using the addaa command. However, I am unable to simulate its secondary structure using ksdssp. I am not sure how to run this command in command line. Or am I doing something wrong in terms of predicting the structures.

Can you please let me know what do I need to do in terms of getting the structure of protein after addsing residues on N-terminus.

Thanks, Haresh

On Tue, Jan 28, 2014 at 2:46 PM, Elaine Meng meng@cgl.ucsf.edu wrote:

No, don't use 9 for the charge of TFLE! Total charge is including the whole protein. Presumably TFLE charge is 0, i.e., it is not a charged sidechain.

Also you should delete hydrogens and then use Surface Color, as in my previous instructions. Elaine

On Jan 28, 2014, at 11:37 AM, Haresh Tukaram More htm211@nyu.edu wrote:

...

Hi Elaine,

Finally!! it worked. In the reply log it shows following

Charges for residue TFLE determined Assigning partial charges to residue CL (net charge -1) with gasteiger

method

...

Total charge for #0: 9.000

So i should use +9 charge for TFLE and TFLE+TFLE? I did that and now

able to get the electrostatic surface potential. The way I did is as follows

...

1. AddH
2. Add charge
3. Select surface of protein
4. Select coulombic surface coloring command and selected the grid

option.

...

5. Next the surface color box opened and I selected color option.
6. Once it is done I removed hydrogen by del H

and I got the protein with electrostatic surface potential.

I hope now this is correct.

Regards, Haresh

On Tue, Jan 28, 2014 at 1:44 PM, Elaine Meng meng@cgl.ucsf.edu wrote: Hi Haresh, If you use "Gasteiger" instead of "AM1-BCC" in the Add Charge dialog as

suggested in my first reply, it works (at least for me, but I'm pretty sure it will for you too)!

...

Elaine

On Jan 28, 2014, at 10:29 AM, Haresh Tukaram More htm211@nyu.edu

wrote:

...

...

HI Elaine,

I tried to do that, but not able to add charges manually. I have

attached a PDB filer here with TFLE mutations. Can you please see if you can add charges on it. I am unable to give charge to TFLE as all the time the process stop at antechamber.exe and abruptly closes the chimera.

...

...

Thanks & Regards, Haresh

On Tue, Jan 28, 2014 at 12:27 PM, Elaine Meng meng@cgl.ucsf.edu

wrote:

...

...

Hi Haresh, There is another issue you may wish to consider: Add Charge will

require adding hydrogens first, but then the molecular surface will be more "craggy" due to the additional small atoms. Here are instructions if you want to show the Coulombic ESP coloring on the less rugged surface calculated without hydrogens (as is usually done; radii automatically adjust to compensate):

...

...

After adding hydrogens and charges: (1) when you use the Coulombic Surface Coloring tool, turn on the

option to "Compute grid..." (in my test just now I took the defaults for the other grid options)

...

...

(2) delete the hydrogens, e.g. command: del H

(3) use the "Surface Color" dialog that automatically appeared when

you created the grid to color the main surface by electrostatic potential using the potential file (grid) you just created. If you don't see that dialog, you can raise it using menu: Tools... Surface/Binding Analysis... Electrostatic Surface Coloring. The default name of that grid is Coulombic ESP. (Do not use the Coulombic Surface Coloring dialog for coloring! After the hydrogens with many positive charges have been deleted, mostly negative charges will be left, and you will get an incorrect very red surface using the Coulombic Surface Coloring dialog.)

...

...

Normally the protein wouldn't have strange residues that require a

separate charge calculation, and this circuitous process wouldn't be required.

...

...

Best, Elaine

---

Elaine C. Meng, Ph.D. UCSF Computer Graphics Lab (Chimera team) and Babbitt Lab Department of Pharmaceutical Chemistry University of California, San Francisco

On Jan 27, 2014, at 6:56 PM, Haresh Tukaram More htm211@nyu.edu

wrote:

...

...

...

Thanks a lot Elaine. I will try to work on it now and see if I can

get any results.

...

...

...

Regards, Haresh

On Mon, Jan 27, 2014 at 12:28 PM, Elaine Meng meng@cgl.ucsf.edu

wrote:

...

...

...

Hi Haresh, The Reply Log message (see the bottom part of what you sent) says

there was a problem calculating the charges of your mutated residues and that you should try running Add Charge first, so that's what I would recommend. Add Charge is in the menu under Tools.. Structure Editing, or can be run with command "addcharge". It has two options for calculating the charges of nonstandard residues: AM1-BCC or Gasteiger. Since the message shows there was a problem calculating using the AM1-BCC method on your mutated residues, I would try the Gasteiger method instead.

...

...

...

<

http://www.rbvi.ucsf.edu/chimera/docs/ContributedSoftware/addcharge/addcharg...

...

...

...

<

http://www.rbvi.ucsf.edu/chimera/docs/UsersGuide/midas/addcharge.html%3E

...

...

...

Then, after you run Add Charge successfully, you can run Coulombic

again. It will automatically detect that the charges have already been calculated, and use those charges for calculating the ESP and coloring the surface.

...

...

...

Actually I tried calculating charges for a single

5,5,5-trifluoroleucine residue by itself (in Chimera, can use "open pubchem:259832" to fetch it by PubChem CID)

...

...

...

<

http://pubchem.ncbi.nlm.nih.gov/summary/summary.cgi?cid=259832&loc=ec_rc...

...

...

...

...and both methods, AM1-BCC and Gasteiger, worked fine; however

maybe the calculation is more difficult in your structure where it sounds like the two mutated residues are treated together.

...

...

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I hope this helps, Elaine

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Elaine C. Meng, Ph.D. UCSF Computer Graphics Lab (Chimera team) and Babbitt Lab Department of Pharmaceutical Chemistry University of California, San Francisco

On Jan 26, 2014, at 8:52 PM, Haresh Tukaram More htm211@nyu.edu

wrote:

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Hi, I tried to get the Coulombic surface coloring for one of the

protein that I modified by removing certain residue at the C-termini and then mutated some residues to the non-natural amino acid 5,5,5-trifluoroleucine using Swisssidechin option. I am getting this error which I am not able to understand.

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Disabled GPU programs because a graphics driver bug was encountered while compiling a vertex shader.

#0, chain A: COMP #0, chain B: COMP #0, chain C: COMP #0, chain D: COMP #0, chain E: COMP C:\Program Files\Chimera 1.8.1\bin\mscalc.exe 1.400000 2.000000 1 MSMSLIB 1.3 started on Local PC Copyright M.F. Sanner (March 2000) Compilation flags

Surface CC TFLE.pdb, category main, probe radius 1.4, vertex

density 2

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22 connected surface components Total solvent excluded surface area = 10393.6 component areas = 8599.46, 624.842, 225.9, 216.153, 81.7059,

54.8894, 54.839, 48.0624, 45.9525, 43.9588, 41.5667, 39.1204, 38.7549, 37.0513, 35.5418, 33.6712, 32.9167, 28.9293, 27.8262, 27.7523, 27.7319, 26.9429

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Total solvent accessible surface area = 10372 component areas = 9971.67, 202.012, 92.2225, 70.7414, 13.8165,

4.12717, 3.62753, 3.24856, 2.597, 1.1866, 1.72654, 1.33137, 1.08737, 0.962375, 0.64812, 0.231422, 0.40193, 0.120068, 0.0570046, 0.0515394, 0.0674059, 0.0311705

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No SEQRES records for combination (#-2147483648.-2147483648) chain

A; guessing terminii instead

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No SEQRES records for combination (#-2147483648.-2147483648) chain

B; guessing terminii instead

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No SEQRES records for combination (#-2147483648.-2147483648) chain

C; guessing terminii instead

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No SEQRES records for combination (#-2147483648.-2147483648) chain

D; guessing terminii instead

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No SEQRES records for combination (#-2147483648.-2147483648) chain

E; guessing terminii instead

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Chain-initial residues that are actual N terminii:

#-2147483648.-2147483648 MET 27.A, #-2147483648.-2147483648 MET 27.B, #-2147483648.-2147483648 MET 27.C, #-2147483648.-2147483648 MET 27.D, #-2147483648.-2147483648 MET 27.E

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Chain-initial residues that are not actual N terminii: Chain-final residues that are actual C terminii:

#-2147483648.-2147483648 TFLE 67.A, #-2147483648.-2147483648 TFLE 67.B, #-2147483648.-2147483648 TFLE 67.C, #-2147483648.-2147483648 TFLE 67.D, #-2147483648.-2147483648 TFLE 67.E

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Chain-final residues that are not actual C terminii: 424 hydrogen bonds Hydrogens added Charge model: AMBER ff12SB Assigning partial charges to residue TFLE+TFLE (net charge +0)

with am1-bcc method

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Running ANTECHAMBER command: C:/Program Files/Chimera

1.8.1/bin/amber12/bin\antechamber -ek qm_theory='AM1', -i c:\users\haresh~1.pol\appdata\local\temp\tmpipssqi\ante.in.mol2 -fi mol2 -o c:\users\haresh~1.pol\appdata\local\temp\tmpipssqi\ante.out.mol2 -fo mol2 -c bcc -nc 0 -j 5 -s 2

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(TFLE+TFLE)

Cannot automatically determine charges for residue TFLE+TFLE; Run AddCharge tool manually to add charges and then rerun ESP

Can someone please tell me a step by step procedure to make this

work.

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Thanks & Regards, Haresh

---

Chimera-users mailing list Chimera-users@cgl.ucsf.edu http://plato.cgl.ucsf.edu/mailman/listinfo/chimera-users

Hi Elaine,

I made two different models in a single Chimera window and selected the N-terminal amino acid of one peptide and C-terminal amino acid of another. Then I ran the command Join mdoels, but it gives me the error. Can you please tell me what I am doing wrong here.

I have attached screen shot of the window.

Thanks, Haresh

On Tue, Jan 28, 2014 at 6:36 PM, Elaine Meng meng@cgl.ucsf.edu wrote:

Hi Haresh, To run ksdssp, just enter command "ksdssp" ... this does not change the coordinates of the structure, it just tries to identify which parts are helix and which are strand based on where it sees backbone H-bonds. If the ribbon is still not like the helix or strand width after you use "ksdssp" it means that the conformation does not look like helix or strand according to ksdssp.

When you use addaa, make sure to specify the desired conformation. http://www.rbvi.ucsf.edu/chimera/docs/UsersGuide/midas/addaa.html

Another way instead of addaa to build the 14 residues is with Build Structure (in menu under Tools... Structure Editing), section Start Structure, choice "peptide", which will then give a further dialog for setting phi/psi angles with suggestions available for different secondary structures. < http://www.rbvi.ucsf.edu/chimera/docs/ContributedSoftware/editing/editing.ht...

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You would build the peptide as a separate new model, but then use the Join Models section of Build Structure to attach it to the N-term of your protein. < http://www.rbvi.ucsf.edu/chimera/docs/ContributedSoftware/editing/editing.ht...

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HOWEVER, even if you built the conformation correctly, it depends whether you tried to make a strand or a helix. Even if the peptide is in a beta-strand conformation, ksdssp will not identify a single strand by itself. It only identifies a beta-strand when the backbone is positioned to H-bond with another beta-strand, as in a hairpin or sheet.

You could just force Chimera to show the residues as helix or strand no matter what the reality is or what ksdssp thinks, for example, selecting all those residues and then if you wanted them to be considered strand, using commands: setattr r isHelix false sel setattr r isStrand true sel (just trade "isHelix" and "isStrand" in those commands if you wanted the residues to be considered helix)

For helix, an additional possibility is to try using ksdssp again but with command-line options to use less strict parameter values. < http://www.rbvi.ucsf.edu/chimera/docs/UsersGuide/midas/ksdssp.html%3E

You can search the manual for strings like "ksdssp" or "building peptides" from the Chimera Help menu. Best, Elaine

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Elaine C. Meng, Ph.D. UCSF Computer Graphics Lab (Chimera team) and Babbitt Lab Department of Pharmaceutical Chemistry University of California, San Francisco

On Jan 28, 2014, at 2:43 PM, Haresh Tukaram More htm211@nyu.edu wrote:

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Hi Elaine, Yes I did exactly the way you explained and it is working now. I have another question. I need to add some 14 residues on N-termius of

the protein which I did using the addaa command. However, I am unable to simulate its secondary structure using ksdssp. I am not sure how to run this command in command line. Or am I doing something wrong in terms of predicting the structures.

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Can you please let me know what do I need to do in terms of getting the

structure of protein after addsing residues on N-terminus.

...

Thanks, Haresh

Hi Haresh, I was able to add charges / ESP color the surface for your structure without any problem on both my Mac and a Windows machine. I will be sending you a session file in a separate mail with the colored surface. On a Windows machine, while the charge calculation is ongoing a second window pops up (with 'antechamber.exe' at the end of its title). Do not close that window. Closing that window will cause the charge calculation to stop/fail. The calculation takes several minutes, so be patient.

--Eric

Eric Pettersen UCSF Computer Graphics Lab http://www.cgl.ucsf.edu

On Jan 28, 2014, at 10:29 AM, Haresh Tukaram More htm211@nyu.edu wrote:

HI Elaine,

I tried to do that, but not able to add charges manually. I have attached a PDB filer here with TFLE mutations. Can you please see if you can add charges on it. I am unable to give charge to TFLE as all the time the process stop at antechamber.exe and abruptly closes the chimera.

Thanks & Regards, Haresh

On Tue, Jan 28, 2014 at 12:27 PM, Elaine Meng meng@cgl.ucsf.edu wrote: Hi Haresh, There is another issue you may wish to consider: Add Charge will require adding hydrogens first, but then the molecular surface will be more "craggy" due to the additional small atoms. Here are instructions if you want to show the Coulombic ESP coloring on the less rugged surface calculated without hydrogens (as is usually done; radii automatically adjust to compensate):

After adding hydrogens and charges: (1) when you use the Coulombic Surface Coloring tool, turn on the option to "Compute grid…" (in my test just now I took the defaults for the other grid options)

(2) delete the hydrogens, e.g. command: del H

(3) use the "Surface Color" dialog that automatically appeared when you created the grid to color the main surface by electrostatic potential using the potential file (grid) you just created. If you don't see that dialog, you can raise it using menu: Tools… Surface/Binding Analysis… Electrostatic Surface Coloring. The default name of that grid is Coulombic ESP. (Do not use the Coulombic Surface Coloring dialog for coloring! After the hydrogens with many positive charges have been deleted, mostly negative charges will be left, and you will get an incorrect very red surface using the Coulombic Surface Coloring dialog.)

Normally the protein wouldn't have strange residues that require a separate charge calculation, and this circuitous process wouldn't be required. Best, Elaine

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Elaine C. Meng, Ph.D. UCSF Computer Graphics Lab (Chimera team) and Babbitt Lab Department of Pharmaceutical Chemistry University of California, San Francisco

On Jan 27, 2014, at 6:56 PM, Haresh Tukaram More htm211@nyu.edu wrote:

...

Thanks a lot Elaine. I will try to work on it now and see if I can get any results.

Regards, Haresh

On Mon, Jan 27, 2014 at 12:28 PM, Elaine Meng meng@cgl.ucsf.edu wrote: Hi Haresh, The Reply Log message (see the bottom part of what you sent) says there was a problem calculating the charges of your mutated residues and that you should try running Add Charge first, so that's what I would recommend. Add Charge is in the menu under Tools.. Structure Editing, or can be run with command "addcharge". It has two options for calculating the charges of nonstandard residues: AM1-BCC or Gasteiger. Since the message shows there was a problem calculating using the AM1-BCC method on your mutated residues, I would try the Gasteiger method instead.

http://www.rbvi.ucsf.edu/chimera/docs/ContributedSoftware/addcharge/addcharge.html http://www.rbvi.ucsf.edu/chimera/docs/UsersGuide/midas/addcharge.html

Then, after you run Add Charge successfully, you can run Coulombic again. It will automatically detect that the charges have already been calculated, and use those charges for calculating the ESP and coloring the surface.

Actually I tried calculating charges for a single 5,5,5-trifluoroleucine residue by itself (in Chimera, can use "open pubchem:259832" to fetch it by PubChem CID) http://pubchem.ncbi.nlm.nih.gov/summary/summary.cgi?cid=259832&loc=ec_rcs ...and both methods, AM1-BCC and Gasteiger, worked fine; however maybe the calculation is more difficult in your structure where it sounds like the two mutated residues are treated together.

I hope this helps, Elaine

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Elaine C. Meng, Ph.D. UCSF Computer Graphics Lab (Chimera team) and Babbitt Lab Department of Pharmaceutical Chemistry University of California, San Francisco

On Jan 26, 2014, at 8:52 PM, Haresh Tukaram More htm211@nyu.edu wrote:

...

Hi, I tried to get the Coulombic surface coloring for one of the protein that I modified by removing certain residue at the C-termini and then mutated some residues to the non-natural amino acid 5,5,5-trifluoroleucine using Swisssidechin option. I am getting this error which I am not able to understand.

Disabled GPU programs because a graphics driver bug was encountered while compiling a vertex shader.

#0, chain A: COMP #0, chain B: COMP #0, chain C: COMP #0, chain D: COMP #0, chain E: COMP C:\Program Files\Chimera 1.8.1\bin\mscalc.exe 1.400000 2.000000 1 MSMSLIB 1.3 started on Local PC Copyright M.F. Sanner (March 2000) Compilation flags

Surface CC TFLE.pdb, category main, probe radius 1.4, vertex density 2 22 connected surface components Total solvent excluded surface area = 10393.6 component areas = 8599.46, 624.842, 225.9, 216.153, 81.7059, 54.8894, 54.839, 48.0624, 45.9525, 43.9588, 41.5667, 39.1204, 38.7549, 37.0513, 35.5418, 33.6712, 32.9167, 28.9293, 27.8262, 27.7523, 27.7319, 26.9429 Total solvent accessible surface area = 10372 component areas = 9971.67, 202.012, 92.2225, 70.7414, 13.8165, 4.12717, 3.62753, 3.24856, 2.597, 1.1866, 1.72654, 1.33137, 1.08737, 0.962375, 0.64812, 0.231422, 0.40193, 0.120068, 0.0570046, 0.0515394, 0.0674059, 0.0311705 No SEQRES records for combination (#-2147483648.-2147483648) chain A; guessing terminii instead No SEQRES records for combination (#-2147483648.-2147483648) chain B; guessing terminii instead No SEQRES records for combination (#-2147483648.-2147483648) chain C; guessing terminii instead No SEQRES records for combination (#-2147483648.-2147483648) chain D; guessing terminii instead No SEQRES records for combination (#-2147483648.-2147483648) chain E; guessing terminii instead Chain-initial residues that are actual N terminii: #-2147483648.-2147483648 MET 27.A, #-2147483648.-2147483648 MET 27.B, #-2147483648.-2147483648 MET 27.C, #-2147483648.-2147483648 MET 27.D, #-2147483648.-2147483648 MET 27.E Chain-initial residues that are not actual N terminii: Chain-final residues that are actual C terminii: #-2147483648.-2147483648 TFLE 67.A, #-2147483648.-2147483648 TFLE 67.B, #-2147483648.-2147483648 TFLE 67.C, #-2147483648.-2147483648 TFLE 67.D, #-2147483648.-2147483648 TFLE 67.E Chain-final residues that are not actual C terminii: 424 hydrogen bonds Hydrogens added Charge model: AMBER ff12SB Assigning partial charges to residue TFLE+TFLE (net charge +0) with am1-bcc method Running ANTECHAMBER command: C:/Program Files/Chimera 1.8.1/bin/amber12/bin\antechamber -ek qm_theory='AM1', -i c:\users\haresh~1.pol\appdata\local\temp\tmpipssqi\ante.in.mol2 -fi mol2 -o c:\users\haresh~1.pol\appdata\local\temp\tmpipssqi\ante.out.mol2 -fo mol2 -c bcc -nc 0 -j 5 -s 2 (TFLE+TFLE)

Cannot automatically determine charges for residue TFLE+TFLE; Run AddCharge tool manually to add charges and then rerun ESP

Can someone please tell me a step by step procedure to make this work. Thanks & Regards, Haresh

---

Chimera-users mailing list Chimera-users@cgl.ucsf.edu http://plato.cgl.ucsf.edu/mailman/listinfo/chimera-users

<TFLE protein.pdb>_______________________________________________ Chimera-users mailing list Chimera-users@cgl.ucsf.edu http://plato.cgl.ucsf.edu/mailman/listinfo/chimera-users