Hello Ana,
it looks very much like a covalent bond with the His. You may get some hints from the chemistry of the reaction, which is normally catalysed. Are there possible side products involving malonate, alternative substrates present in the buffer etc ?
Occupancy refinements of the different atom groups may also yield
some information.
This looks really interesting
Cheers
Wim
I
Hello Jon,Thanks for the comments. Yes, the His is part of the active site, and the distances are very close, like a covalent bond. That's why I was wondering if the malonate can be bound to the His.
We tried to model the malonate in the different conformation, but that didn't work. But I'll check about the protease inhibitor. Thanks!
Kind regardsAna
On Tue, 17 Aug 2021 at 14:48, Jon Cooper <jon.b.coo...@protonmail.com> wrote:
Hello, only other thought was that malonate might be binding in two conformations, i.e. dual occupancy, and did you use a protease inhibitor cocktail, since the constituents can react? The density looks a bit like citrate, but too close to the His. I couldn't read the distances to the His in your figure. Is it part of the active site and if so can you say what the enzyme is? Anomalous difference maps are popular here, too. Is the surrounding structure totally OK because odd features in the difference map can indicate problems nearby e.g. I did see a leucine which looked slightly strained on the left, but maybe I am wrong. Good luck. Cheers, Jon.C.
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-------- Original Message --------
On 17 Aug 2021, 12:11, Ana Ebrecht < anaebre...@gmail.com> wrote:
Hi Jon,Thanks for the reply. I don't think this is acetylation, because I only see this density in the crystals that grew with malonate. In other conditions doesn't show anything like that. So I thought it'd be the malonate. But I'll check on that.Thanks for the suggestion.
Kind regardsAna
On Mon, 16 Aug 2021 at 16:56, Jon Cooper <jon.b.coo...@protonmail.com> wrote:
Hello, could the His be partially acetylated?
Best wishes Jon.C.
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-------- Original Message --------
On 16 Aug 2021, 14:52, Ana Ebrecht < anaebre...@gmail.com> wrote:
Dear all,
I am building the structure of a protein that was crystallized in 0.2 M sodium malonate pH 5.0, 20% w/v polyethylene glycol 3,350.During the refinement, we found what we think is a malonate molecule in the active site, but it seems like is bound somehow to the histidine (this His is the catalytic residue of the enzyme), almost like a covalent interaction. Under other conditions of crystallization, the protein bound a sulfate and an acetate in the site but did not show this type of interaction with the histidine.
We couldn't find anything that explains a reaction between the malonate and the histidine.
Does anyone have experience with this reaction or have seen something similar before?
ThanksKind regardsAna
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