Hi Hiroshi,

  I think everyone agrees that you cannot trust AlphaFold or other computational structure prediction methods to decide if proteins bind.  These computational predictions are just to give you clues about the possible behavior and must be tested experimentally.  In the case of gp64 AlphaFold 3, Boltz 2, OpenFold 3, and AlphaFold 2 all have low confidence that it forms a dimer.  But if experiments show it forms a dimer then quite possibly the computational predictions are wrong.  Of course you have to just the weight and reliability of experimental evidence also.  I am not an experimentalist and do not know how reliable an SDS-PAGE gel is at identifying oligomeric protein states.

  The computational predictions are not for "proteins moving freely in solution".  These computational programs know nothing about water or dynamics of proteins.  They were trained on experimental structures from the Protein DataBank which are mostly derived from X-ray crystallography and electron cryo-microscopy, and those techniques have the proteins frozen in non-crystalline ice.

  Also as you point out gp64 is attached to membranes, and the membrane environment may be necessary for them to dimerize.

  In summary, the computational predictions do not provide decisive answers about protein-protein interactions.

Tom


On Jul 8, 2026, at 5:25 PM, 落合廣 via ChimeraX-users <chimerax-users@cgl.ucsf.edu> wrote:

Hi Dr, 
Thank you very much for conducting the analysis using various versions of AlphaFold.  Based on the AlphaFold criteria, the conclusion appears to be that gp64 is unlikely to form a proper dimer. While I may have some misunderstandings due to my limited expertise in using AlphaFold and ChimeraX,  I believe that—considering the following points—it cannot be definitively ruled out that gp64 forms a dimer. Specifically, AlphaFold3 predicts the structure as a "protein moving freely in solution," whereas in reality, gp64 is anchored to the membrane surface via a GPI anchor.
1. A dimer-like band is observed in SDS-PAGE.
2. It contains a Sushi domain (characterized by four disulfide bridges); Sushi domains are known to be involved in protein-protein associations (as seen in "complement-containing proteins").
3. ChimeraX analysis indicates an interface involving approximately 58 residues.
4. The interface surface area is 1,398 Ų.
Given these factors, I would appreciate your opinion on how this should be interpreted.
with best regards,

 

Hiroshi Ochiai

2026/07/05 14:29、落合廣 <ochiai3@icloud.com>のメール:

Dear DR.
Thank you for using chimeraX. I am very impressed with the development of protein assembly analysis technology. I am an elderly (88-year-old) biochemist.
　I have a question.
○1. Even if dimer formation is analyzed by AlphaFold 3, dimers with interfaces are not recognized at all (AlphaFold has a list of 19(?) analyzed). Why?
○2. Even though dimer formation (54 or 132 interface residues) can be seen by chimeraX, when looking at ipTM with AlphaFold, a number such as 0.19 is not recognized as a dimer. Do you think that this is because AlphaFold does not take into account that proteins such as GPI membrane proteins are kept in anti-parallel, do you have an opinion?
With best Regards

Hiroshi Ochiai
Research Institute for Electronic Sciences, Hokkaido University, 
Sapporo Japan
Emeritus Professor

_______________________________________________
ChimeraX-users mailing list -- chimerax-users@cgl.ucsf.edu
To unsubscribe send an email to chimerax-users-leave@cgl.ucsf.edu
Archives: https://mail.cgl.ucsf.edu/mailman/archives/list/chimerax-users@cgl.ucsf.edu/