[Myotox] toxin/lipid interactions

Tue Dec 7 09:27:14 MST 2021

Dear Bruno,
thanks for getting back - since it is essential that Bruno is part of
this discussion I will try once more with a new poll for next week:

https://doodle.com/poll/ut3psa4k68tiu6ve?utm_source=poll&utm_medium=link

Please check your browsers, I believe the times listed are for Mountain
time, for comparison, Mondays options are 11:00 am - 4:00 pm, 1 hour
sections.

Please all respond by the end of the day so we can release the other 
times for everyone. I will send out the consensus by tonight.

Thanks, -Borries

On Mon, Dec 06, 2021 at 10:27:49PM -0600, Bruno Lomonte wrote:
> Dear Borries and All,
> 
> Many thanks Borries for envisioning all these interesting avenues to study
> and better understand the toxin. It will be exciting to explore the
> possibility of pore formation or other mechanisms related to myotoxicity.
> There has been a large knowledge gap regarding the interaction of this type
> of toxin with membranes, so this is much needed work!
> 
> I olny regret at this moment that the following days (proposed in the Doodle
> poll) are fully booked in my case. Sorry! If there are other alternative
> options for everyone to meet and discuss, please let us know.
> 
> Here attached is an old study done on liposome permeabilization (including
> liposomes made of non-hydrolyzable ether lipids - the toxin can disrupt
> them, but nothing further has been done along this line of work, as far as I
> know.
> 
> Best regards,
> 
> Bruno
> 
> ++++++++++++++++++++++
> 
> 
> On 12/6/2021 12:00 PM, Borries Demeler wrote:
> > Hello everybody,
> > 
> > it has been a while since we have reported back on our NMR findings,
> > so it is about time to pick up this interesting project again and get
> > everyone up-to-date on the latest status.
> > 
> > If you recall, Amy found that by using significantly lower SDS
> > concentrations than CMC that myotoxin-II formed a very stable and
> > homogeneous oligomer with about 6 or 7 myotoxin-II molecules in it. When
> > SDS concentration was increased, it was possible to disrupt this oligomer
> > and generate increasing amounts of heterogeneous oligomeric forms, with
> > the predominant form being dimer. This observation matched nicely with
> > the SDS PAGE results reported earlier by Susumu Uchiyama's group.
> > 
> > But of course, more interesting to all of us should be what the
> > structure of the oligomeric species was, and if it potentially formed
> > a membrane penetrating pore, or has some other functional relevance.
> > 
> > If you recall, and in summary, Paul and his group has investigated the
> > interaction of SDS with myotoxin-II, and after multiple iterations of
> > different ratios of SDS to myotoxin-II, concluded the following:
> > 
> >     "There seems to be an unusually strong interaction with SDS. The
> >     alpha methylene exhibits high stress similar to that seen in an epoxy
> >     ring. Ar first sight it appears to be an AB quartet. Simulations show
> >     that the 2JHH of the methylene is very small indicating strain."
> > 
> > Paul also mentioned that the NMR studies are hindered by the large
> > background signals and really would require labeling to allow more
> > sophisticated multi-dimensional analysis.
> > 
> > I also pursued the potential to get more info with cryo-EM by sending
> > the samples to the University of Manitoba for study by Pauline Padilla
> > and Joerg Stetefeld, but this angle unfortunately did not go anywhere,
> > since negative staining screens did not produce the desired results,
> > suggesting cryoEM is out of the question.
> > 
> > Separately, I contacted Bruce Bowler at the University of Montana, who
> > studies protein-membrane interactions, and he offered an array of other
> > lipids he had used for crystallizing cytochrome-C interacting with
> > membranes and still has in the fridge. So we could try to use some of
> > these lipids to replicate Amy's results with other lipid systems.
> > 
> > Recently, I heard a very interesting talk by Sebastien Poget on the
> > interactions of various animal toxins with lipids, so I shared our
> > findings with him. He wrote the following:
> > 
> >     "Thanks for your e-mails, it looks like you have a very interesting
> >     toxin to study. Is there any precedent for a snake phospholipase
> >     forming membrane channels? It would make sense physiologically,
> >     as a quick way to disrupt the membrane and place the phospholipase
> >     in the right environment to hydrolyze the lipids. One way that
> >     you could quickly look into this would be to add the phospholipase
> >     to artificial bilayers - one could maybe use ether-linked lipids
> >     to prevent hydrolysis and see if any pores are formed just by the
> >     presence of the toxin alone. We have a planar bilayer system in my lab,
> >     and I would be happy to give this a try if it would be of interest to
> >     you. As you mentioned, testing oligomerization in the presence of other
> >     detergents may also be interesting, including maybe lysolipids. Maybe
> >     we can set up a zoom call to discuss some of these questions in more
> >     detail if you are interested?  Best, Sebastien"
> > 
> > I am passing his questions on to this group to get your comments.
> > Sebastien is now included on our email list, so please feel free to
> > comment by simply replying to this message. I personally would be very
> > curious to see if such a membrane pore theory could be tested with
> > Sebastien's methods.
> > 
> > I would like to follow Sebastien's suggestion and propose a zoom call to
> > discuss next steps. Here is a Doodle poll for a zoom meeting, please
> > respond by the end of tomorrow.
> > 
> > https://doodle.com/poll/xbx5rhhf9g7mkxb3?utm_source=poll&utm_medium=link
> > 
> > If none of the proposed times work for most of us, we could try again
> > next week with another doodle poll.
> > 
> > Thanks! -Borries
> > _______________________________________________
> > Myotox mailing list
> > Myotox at biophysics.uleth.ca
> > https://demeler7.uleth.ca/mailman/listinfo/myotox
> 
> -- 
> Bruno Lomonte, Ph.D.
> Instituto Clodomiro Picado
> Facultad de Microbiología
> Universidad de Costa Rica
> San José, COSTA RICA
> 
> tel.of. (+506) 2511 7888
> bruno.lomonte at ucr.ac.cr

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