locked nucleic acids/fiber

[amirtaghavi]

Hello,

I have a question regarding the fiber option of 3DNA. Is it possible to add standard locked nucleotides or user modified ones to the fiber data base so one can create RNA structures which is a mix of standard and locked nucleotides. 
For example I have a regular sequence of "AACGU" and I wand to change the second "A" to "LA", assuming I have the pdb information of "LA".
Thanks in forward for your help.

Best regards,
Amir

[xiangjun]

Hi Amir,

It is possible as you suggested, although there are other ways using 3DNA. I may be able to help if you're specific on what you want to achieve.

Best regards,

Xiang-Jun

[amirtaghavi]

Hi Prof. Xiang-Jun,

Thanks a lot for the reply. So I have a modified locked nucleotide named N7-LNA-adenosine and I want to make the following sequence:
UCAG"7AL"CAGU (I have the geometry optimized pdb file of the 7AL if it is required).
Thanks for your help.

Best regards,

[xiangjun]

What 7AL looks like? Please attach an example PDB file. Do you want to create a single-stranded RNA structure with sequence UCAGaCAGU ('a' for 7AL), right? If so, then please use the 3DNA 'fiber' program to generate a canonical RNA model. We can start from there.

As a general note, it is always helpful to be concrete, and provide as much info as possible. Guessing usually does not help much.

Xiang-Jun

[amirtaghavi]

Please find attached the pdb file of 7AL. The main changes are that sugar is attached to N7 (instead of N9) and there is a bridge connection between C2', C3' and C4' in the sugar.
I want to make a double stranded RNA which in one strand "A" would be replaced by 7AL ('a') ,UCAGaCAGU, but the second strand remains intact.
Using fiber I have made the dsRNA with the mentioned sequence.

Thanks a lot.

[xiangjun]

Thanks for providing the following two crucial clarifications:

• So "7AL" has a sugar attached to N7 (instead of N9). The attached PDB file makes it unambiguous.
• You want to 'make a double stranded RNA which in one strand "A" would be replaced by 7AL'.

With these details, I'd say that 3DNA cannot fit your direct needs completely. You may well want to try other advanced software tools with an interactive interface (e.g., Maestro from Schrödinger). Nevertheless, the following steps may help you in the right direction. If you still want to give DSSR a try, please proceed as suggested and report back your results.

Code: Bash

1. # set 7AL in the standard base reference frame, assuming N9--C1' connection
2. std_base -A -fit 7AL.pdb 7AL-ref-fit.pdb
3. # rotated the above '7AL-ref-fit.pdb' file by y-axis of 180 degrees because of N7--C1' linkage
4. echo 'by rotation y 180' > roty180
5. rotate_mol -r=roty180 7AL-ref-fit.pdb 7AL-refOK-fit.pdb
6. # generate the duplex RNA fiber model with proper sequence
7. fiber -rna -seq=UCAGACAGU RNA-fiber.pdb
8. # get the file with base reference frames in 'ref_frames.dat'
9. find_pair -s RNA-fiber.pdb temp.txt
10. # reorient the above fiber model in the reference frame of A5
11. frame_mol -5 ref_frames.dat RNA-fiber.pdb RNA-fiber-ref5.pdb
12. # load 7AL-refOK-fit.pdb and RNA-fiber-ref5.pdb into PyMOL

HTH,

Xiang-Jun

[amirtaghavi]

Thanks a lot for your help and time. I am trying to use the combination of chimera and xleap (amber) to make the structure as I don't have access to maestro. I will also try DSSR see if I can make it work (thanks for the script). I appreciate your help and time.

Best regards,

[amirtaghavi]

Dear Prof. Xiang-Jun,

So, I made a small change in your script so that I could  remove the nucleotide ('A') which I wanted to replace with 7AL and then loaded both structures (7AL-refOK-fit.pdb and RNA-fiber-ref5.pdb) in pymol and saved it as a single file.
then I loaded the pdb file in xleap (as I have already added the 7AL parameters to the library) and the missing bond was fixed to get the final structure.
your script was very helpful and I learned a lot. I wish there was a few such scripts which could help to better understand the powerful tools of 3DNA.   
again thanks a lot for your help.

Cheers,

[xiangjun]

Glad to hear that the 3DNA-command script helped in your case.

If you have further questions that are broadly related to 3DNA (or DSSR and SNAP), please do not hesitate to ask on the Forum. In asking questions, please be as specific as possible so others can reproduce the issue. It also helps that a follow-up summary post is provided by the initiator of the thread.

Best regards,

Xiang-Jun