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preeti Offline OP
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Dear All,

I am experiencing problems ith reading phosphorylated residue THR which is at position 309 in the original pdb file. I did the following:

-- First I removed TPO residue from the pdb file and made a separate pdb (attached - at the end of this message).
-- Second, I renumbered pdb (attached- "testa-renum.pdb.gz") containing all protein residues expect tpo to make numbers continuous.

However, I get the following error using c33b2 version of charmm. Can anyone help me with this problem?
________________________________________________________
CHARMM> OPEN READ UNIT 22 CARD NAME tpo.pdb
VOPEN> Attempting to open::tpo.pdb::
OPNLGU> Unit 22 opened for READONLY access to tpo.pdb

CHARMM> READ SEQU PDB UNIT 22
MAINIO> Sequence information being read from unit 22.
TITLE> *

RESIDUE SEQUENCE -- 1 RESIDUES
TPO

CHARMM> generate TPO SETUP WARN

***** ERROR in GENIC ***** Residue 'TPO ' was not found.
________________________________________________________

INPUT FILE:

* STARTING WITH PDB FILE ...
* THIS IS phosphorylated THR structure
*

open unit 1 read form name top_all27_prot_na.rtf
read rtf unit 1 card
close unit 1

open unit 1 read form name par_all27_prot_na.prm
read param unit 1 card
close unit 1

bomlev -1

stream toppar_prot_na_all.str

set initresa 146
set one 1
calc offseta = (-@one*@initresa) + @one

open unit 1 read card name testa-renum.pdb
read sequ pdb unit 1
generate AKTA setup warn
rewind unit 1

OPEN READ UNIT 22 CARD NAME tpo.pdb
READ SEQU PDB UNIT 22
generate TPO SETUP WARN

AUTOGENERATE ANGLES DIHEDRALS
rewind unit 22

!LINK C LYS A 308 N TPO A 1
!LINK C TPO A 1 N PHE A 309

patch THP2 TPO AKTA 308 SETUP WARN
patch THP2 TPO AKTA 309 SETUP WARN

read coor unit 1 pdb offset @offseta append
close unit 1

READ coor PDB UNIT 22
CLOSE UNIT 22

stop

___________________
TPO.pdb is as following:

ATOM 1 N TPO A 1 22.995 22.753 176.816 1.00 14.80 TPO
ATOM 2 CA TPO A 1 24.145 22.248 176.051 1.00 12.61 TPO
ATOM 3 CB TPO A 1 24.836 23.392 175.261 1.00 13.99 TPO
ATOM 4 CG2 TPO A 1 25.863 22.833 174.267 1.00 11.74 TPO
ATOM 5 OG1 TPO A 1 23.864 24.142 174.505 1.00 13.97 TPO
ATOM 6 P TPO A 1 23.695 25.504 174.704 1.00 15.80 TPO
ATOM 7 O1P TPO A 1 24.865 26.333 174.283 1.00 15.57 TPO
ATOM 8 O2P TPO A 1 23.252 25.585 176.140 1.00 15.91 TPO
ATOM 9 O3P TPO A 1 22.569 25.829 173.656 1.00 15.73 TPO
ATOM 10 C TPO A 1 25.200 21.515 176.902 1.00 13.83 TPO
ATOM 11 O TPO A 1 25.651 22.046 177.914 1.00 12.56 TPO

Attached Images
19136-testa-renum.pdb (0 Bytes, 275 downloads)
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rmv Online Content
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Unless you've added a TPO residue description to the topology file, I'd expect that error.

I generally try to avoid renumbering and preserve the PBD RESIDs if at all possible, as it makes life much easier.

I suggest either creating TPO from THR, or simply renaming the residue to THR; for the latter case, you'd need a patch that converts the hydroxyl to a phosphate. With a bit of careful study, it should not be too difficult to construct a patch (PRES) that adds a phosphate to SER or THR, assuming transferability from lipid or nucleic acid parameters. Maybe somebody already has one, but I did not find e.g. a phosphoserine patch in my quick look through the toppar subdir.

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preeti Offline OP
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Thanks Rick. I will try.

I found a link below mentioning phosphoserine. I tried the example mentioned and it didn't work either.

Best,
Preeti

http://www.charmm.org/ubbthreads/showflat.php?Cat=0&Number=15487&Main=15399

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rmv Online Content
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That stream file has the needed patch to convert THR to the phosphorylated form. To use it, simply change the TPO residue to THR in the PDB (w/o renumbering!), then read the protein sequence and use GENERate to make the protein segment as usual. Follow that with a PATCH command, e.g. (assuming segid A for the protein, resid 309 for THR):

PATCH THP1 A 309
AUTOGEN ANGLE DIHE

Simply stating that something "didn't work" is usually not enough information to resolve a problem.

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preeti Offline OP
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Hi Rick,

many many thanks for your help. It works and i viewed the odb in vmd and it's perfect.

In the pdb structure, residues 450 to 465 are missing (disordered). I didn't model the missing residues, but just followed your earlier posts to generate the crd file by joining the two segments after reading them separately from the two pdb files and it worked too in terms of writing the final coordinate file.

My question is: In general, if I want to run dynamics, and explore the conformations of this structure, should I try to model the missing region as a loop or is it acceptable just to assume that missing region has no structure and hence will not be modelled in simulation?

Preeti

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rmv Online Content
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If it were my project, I'd probably look into building coordinates for the disordered loop region. It is probably flexible, but perhaps not unstructured; the missing crystal data implies several possible folds, but not necessarily a lack of structure. It's a question that simulations might help to answer.


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