[Myotox] toxin/lipid interactions

Tue Dec 7 10:02:48 MST 2021

Hi Borries,
Thank you for the detailed update and for the news regarding new
collaborations/opportunities, it will be a pleasure to continue working
with you in this challenging line of research.
Best regards,
Ana

PS.: 11 am in Lethbridge would correspond to 3 pm in Brazil (São Paulo
time), we are 4 hours ahead of you. But the first time option on Doodle
Monday is 16 h (São Paulo time), I got confused.
Anyway, I would only be available on Tuesday afternoon/night (whatever time
it is).

Em ter., 7 de dez. de 2021 às 13:28, Borries Demeler <demeler at gmail.com>
escreveu:

> 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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> > Myotox at biophysics.uleth.ca
> > https://demeler7.uleth.ca/mailman/listinfo/myotox
>
> _______________________________________________
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> Myotox at biophysics.uleth.ca
> https://demeler7.uleth.ca/mailman/listinfo/myotox
>

-- 
Ana Gisele da C. Neves Ferreira
Pesquisadora Titular em Saúde Pública
Chefe do Laboratorio de Toxinologia
Instituto Oswaldo Cruz - Fiocruz
Pavilhao Ozorio de Almeida, sala 05
Av. Brasil, 4365 - Manguinhos
Rio de Janeiro - RJ
21040-900 Brasil
(+5521) 2562-1381 / 98801-5726
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