The following two tutorials show how to use Memoir to model transmembrane proteins with multiple chains. The free PyMOL visualisation software is required for this tutorial. All the sequence and structure files used and all the Memoir generated models are available for download.


1. Modelling a multi-chain ion channel

In the first example we are going to model a homo-oligomer, ion-channel target (PDB: 2WLJ). The full channel is a tetramer made from four copies of the following sequence:

>2WLJ:A|PDBID|CHAIN|SEQUENCE
MTGGMKPPARKPRILNSDGSSNITRLGLEKRGWLDDHYHDLLTVSWPVFITLITGLYLVTNALFALAYLACGDVIENARP
GSFTDAFFFSVQTMATIGYGKLIPIGPLANTLVTLEALCGMLGLAVAASLIYARFTRPTAGVLFSSRMVISDFEGKPTLM
MRLANLRIEQIIEADVHLVLVRSEISQEGMVFRRFHDLTLTRSRSPIFSLSWTVMHPIDHHSPIYGETDETLRNSHSEFL
VLFTGHHEAFAQNVHARHAYSCDEIIWGGHFVDVFTTLPDGRRALDLGKFHEIAQHHHHHH

To model this protein we are going to use PDB structure 3SYO as a template. This structure has a single chain which is rotated to produce a tetramer in the biological unit. In this example we will construct the biological unit from the crystal asymmetric unit. This is possible because PyMOL is able to interpret the CRYST1, ORIGXn, and SCALEn records present in the 3SYO PDB file.

1
Load the 3SYO.pdb in PyMOL. You can either do this via the File → Open menu to load the structure file you downloaded from the PDB or by entering 3SYO in the Plugin → PDB Loader Service menu option. When you first load this, the protein will be first displayed as lines. For a less cluttered cartoon representation click on S → Cartoon and on H → Lines. The S and H menus are found on the right hand side in the below screenshot. The result should look similar to the following image.
3syo loaded
2
In the case of this protein, the biological unit may be reconstructed from the crystal asymmetric unit by issuing the following PyMOL command:
symexp sym,3SYO,(3SYO),1
at the PyMOL> prompt at the bottom of your PyMOL window. Note that each symmetric unit is now a separate PyMOL object (i.e. sym01000000, sym02000000, and sym03000000).
symettric units
3
Save each chain as a separate molecule through the File → Save Molecule menu option. At the end of this exercise you should have four PDB files: 3SYO.pdb, sym01000000.pdb, sym02000000.pdb, and sym03000000.pdb.
4
We now have the inputs we need to run Memoir. Open four instances of Memoir (in different browser tabs). Copy and paste the sequence above in the sequence text box of each browser tab. Then load each of the four PDB files you just created in each different instance of Memoir. The chain should be set to 'A' for all the Memoir runs. A single run should look like the screenshot below. Click on the "Model" button for each Memoir instance. This step will take about 30 minutes to finish.
example memoir submission form
5
When you get the results page, download each "High Accuracy Model" and save each file separately. Load the four models in PyMOL (convert to "cartoon" representation as described in Step 1). Note that using Memoir you can only model the transmembrane region.
load memoir models in PyMOL
6
Save everything as one model using the following PyMOL command at the PyMOL> prompt:
save memoir_2wlj_complete_final.pdb, all

2. Modelling a large cytochrome complex

In the second example we are going to model a large cytochrome complex (PDB: 1L0L). This complex has 11 chains, but we cannot model all of them because they do not all match the template we selected. Also, Memoir does not model non-transmembrane parts such as chain I in this particular example. The three chains we want to model are C, D, and E. The three sequences in FASTA format are given below:

>1L0L:C|PDBID|CHAIN|SEQUENCE
MTNIRKSHPLMKIVNNAFIDLPAPSNISSWWNFGSLLGICLILQILTGLFLAMHYTSDTTTAFSSVTHICRDVNYGWIIR
YMHANGASMFFICLYMHVGRGLYYGSYTFLETWNIGVILLLTVMATAFMGYVLPWGQMSFWGATVITNLLSAIPYIGTNL
VEWIWGGFSVDKATLTRFFAFHFILPFIIMAIAMVHLLFLHETGSNNPTGISSDVDKIPFHPYYTIKDILGALLLILALM
LLVLFAPDLLGDPDNYTPANPLNTPPHIKPEWYFLFAYAILRSIPNKLGGVLALAFSILILALIPLLHTSKQRSMMFRPL
SQCLFWALVADLLTLTWIGGQPVEHPYITIGQLASVLYFLLILVLMPTAGTIENKLLKW
>1L0L:D|PDBID|CHAIN|SEQUENCE
SDLELHPPSYPWSHRGLLSSLDHTSIRRGFQVYKQVCSSCHSMDYVAYRHLVGVCYTEDEAKALAEEVEVQDGPNEDGEM
FMRPGKLSDYFPKPYPNPEAARAANNGALPPDLSYIVRARHGGEDYVFSLLTGYCEPPTGVSLREGLYFNPYFPGQAIGM
APPIYNEVLEFDDGTPATMSQVAKDVCTFLRWAAEPEHDHRKRMGLKMLLMMGLLLPLVYAMKRHKWSVLKSRKLAYRPP
K
>1L0L:E|PDBID|CHAIN|SEQUENCE
SHTDIKVPDFSDYRRPEVLDSTKSSKESSEARKGFSYLVTATTTVGVAYAAKNVVSQFVSSMSASADVLAMSKIEIKLSD
IPEGKNMAFKWRGKPLFVRHRTKKEIDQEAAVEVSQLRDPQHDLERVKKPEWVILIGVCTHLGCVPIANAGDFGGYYCPC
HGSHYDASGRIRKGPAPLNLEVPSYEFTSDDMVIVG

Each of the three chains we wish to model is present in two copies in the biological unit. As a template we are going to use PDB structure 2FYN. We will require 6 Memoir runs to model our target chains — one for each copy of each chain:

  • 1L0L:C with templates 2FYN:A and 2FYN:D
  • 1L0L:D with templates 2FYN:B and 2FYN:E
  • 1L0L:E with templates 2FYN:C and 2FYN:F
1
Launch Memoir in six different browser tabs. In the first two tabs copy and paste sequence 1L0L:C given above as the target we want to model. Then enter 2FYN in the PDB code text box and enter chains A and D in the two different tabs (this should be the only difference between the two tabs). In the second two tabs copy and paste sequence for 1L0L:D given above as the target we want to model, enter 2FYN in the PDB code text box and enter chains B and E in the two different text boxes in the two tabs. Then repeat this for the last two Memoir tabs, i.e. copy and paste sequence for 1L0L:E given above as the target we want to model, enter 2FYN in the PDB code text box and enter chains C and F in the two different tabs. Press the "Model" button in each of the six tabs. An example of how one of these Memoir instances should look is shown below.
example memoir submission form
2
When you get the six results pages, download each "High Accuracy Model" and save each file separately. Load the six models in PyMOL (convert to "cartoon" representation as described in the previous example). Note that using Memoir you can only model the transmembrane region.
load memoir models in PyMOL
3
Save everything as one model using the following PyMOL command:
save memoir_1l0l_complete_final.pdb, all

The above two examples show how to model multiple chain transmembrane proteins using Memoir.