Anti-Infectives Celebration Mur Ligase Meeting May 2024

[Yuhang-CADD]

Date: May 14th 2024
Time: 2pm UK time other timezones

Place: https://ucl.zoom.us/j/91379419977

Recording: https://youtu.be/aFkvye5Vky8?si=cudBQu2f6DDRO77j

Previous Meeting: #109
Who can come?: Anyone. No need to say anything unless you'd like to.

Apologies: Laura, Chris...

Decks: Please @opensourceantibiotics/murligase remember that if you share slides/info, to drag and drop those into a comment on this page, below. Very easy and saves @mattodd having to pester you.

Agenda:

Key things today are:

i) Multi-targeting compounds identified from Warwick and Atomwise collections. Updates on any new IC50 or crystallographic or microbiology data.

ii) New data from Warwick on the contributed AZ compounds (retest at a lower concentration).

iii) Update on crystallographic data of AZ compound 4 (WYH9 compound) in Pae MurD ligase.

(please add if you'd like to prioritise anything)

1) Multi-targeting Compounds from the Warwick/NEU Enamine Collection and Atomwise Compounds

• @AJLloyd105 to update on further enzymatic screening of compounds from the Warwick Enamine Collection and Atomwise libraries, following discovery of the hits shown here.
• @LauraDS1 to update on any soaking and/or co-crystallisation experiments with these compounds, following (last update here.
  Update on crystallography trials at The University of Kansas (Scott Lovell, via @bartrum): Compound J06 (Z2010503124).
• @gfsfernandes to update on current research in design of improved binders vs TB murs.

2) Multi-targeting Elaborated Fragments from Diamond Fragment Screen

• @LauraDS1 to update on screening (inhibition, SPR, soak) of elaborated Diamond fragments sent by @edwintse in Compound to Warwick Inhibition Assay Jun 2022 #84.

3) Variants of AZ Compounds

These will include guanidinium and pyridinium derivatives to try to combat efflux.

• @Yuhang-CADD's 1st batch of fresh samples of AZ5595 and derivatives (Alcohols & Amines) need complete data from @AJLloyd105
• @Yuhang-CADD's 2nd batch of fresh samples of AZ5595 and derivatives (Amines & Guanidiniums) need complete data from @AJLloyd105
• @Yuhang-CADD's 3rd batch of fresh samples of AZ5595 and derivatives (Guanidiniums & Pyridiniums) in progress.

4) New Protein Structures

No new structures to add to the PDB list.

5) De Novo Computational Modelling

• @mattodd to organise what is known of the competition compounds and what is needed to write the work up for publication.

@Yuhang-CADD has paused synthesis of @jhjensen2's originally-suggested compounds (last update was here while the AZ derivatives are being made.

6) CC4CARB Proposal (#93)

• @mattodd and @eyermanncj to finalise current CC4CARB draft and submit. Help is needed from Warwick on commitment to compound evaluation.

7) Misc/AOB

Last time @AJLloyd105 reported on the evaluation, enzymatically, of "double-headed" compounds containing uridine and ATP binders: . These were found to be binders and will be investigated further (need to upload deck). @AJLloyd105 to update as appropriate.

8) Mothballs (if no actions then these need to be linked in wiki and closed)

@mattodd is in the process of converting old items to the wiki

Next Meeting

June 11th 2pm UK time

L'esprit de l'escalier

If you'd like to follow up after the meeting, please comment below. You can also email, but please be clear if anything in the email should not be public domain - the default is always open.

[Yuhang-CADD]

Adrain's report on Atomise
080524.pdf

[AJLloyd105]

080524-140524.pdf
Warwick data

[AJLloyd105]

Hi, Apologies ! Best Wishes, Adrian Adrian Lloyd. Associate Professor, School of Life Sciences, University of Warwick, Gibbet Hill Road, Coventry, CV4 7AL. Tel: Lab +44 (0)2476 522568; Office +44 (0)2476 522700 From: Mat Todd ***@***.***> Sent: Tuesday, May 14, 2024 11:12 PM To: opensourceantibiotics/murligase ***@***.***> Cc: Lloyd, Adrian ***@***.***>; State change ***@***.***> Subject: Re: [opensourceantibiotics/murligase] Anti-Infectives Celebration Mur Ligase Meeting May 2024 (Issue #110) Reopened #110<#110>. - Reply to this email directly, view it on GitHub<#110 (comment)>, or unsubscribe<https://github.com/notifications/unsubscribe-auth/AY5MUXG7LTRUZ3F5AOYRSHTZCKD2TAVCNFSM6AAAAABHUCJCQOVHI2DSMVQWIX3LMV45UABCJFZXG5LFIV3GK3TUJZXXI2LGNFRWC5DJN5XDWMJSHAYDQNZXGMYDANQ>. You are receiving this because you modified the open/close state.Message ID: ***@***.******@***.***>>

[Yuhang-CADD]

Dear @AJLloyd105 and team,

Hi, I hope you are doing well!

Following up with the data that Adrian have presented from last meeting (much appreciate your efforts), I have had some analysis to discuss about here and I would like to have your expert thoughts!

The 1st and 2nd round of AZ derivatives (WYH compounds) are as follows with IC50 and Ki values.

As we can tell from the first two lines, the original alcohol version of the AZ structures was very potent, but the in situ modifications only made these structures less potent by 4 folds from alcohol to amine, and amine to guanidinium.

This somehow matched with the original reference paper (entryway improved rules with guanidinium & pyridinium https://pubs.acs.org/doi/full/10.1021/acsinfecdis.0c00715#): the MIC values against porin-deleted mutants (ΔtolC) decreased as amines were changed to guanidiniums.

(this picture from Hergenrother ACS 2021 was referenced for fair purpose_academic discussions)

However, I am also curious about other factors that may influence the inhibition data this time; for example, whether the actual testing pH (as well as the general conditions) for the potency results of these protonated compounds would matter (despite the fact that pH was more acknowledged as an factor in compound accumulation which will be observable in MICs)?

According to Adrian’s report from last meeting, there might be a pH-dependent sensitivity in MurC with these compounds and the pH condition Adrian applied was at 7.6.

I have had a revisit of relevant hergenrother papers (https://www.nature.com/articles/nature22308#additional-information; https://pubs.acs.org/doi/full/10.1021/acsinfecdis.0c00715#; https://www.nature.com/articles/s41564-019-0604-5)

and I have managed to find the following conditions they used for amines in the inhibition assay (even though it was applied against Fabl enzyme). I am not sure if this pH = 7.8 would help change things around or not? Would be very excited to see the actual MICs of WYH compounds to further confirm.

Interestingly, on the last line of the AZ inhibition results, the WYH30-X-P amine, which was designed to be inactive (expected clashes with MurC C-terminus) got an improvement after the guanidinylation (WYH76-X-P) by 4 folds.

My understanding would be: we might want to carefully monitor whether in our AZ system, guanidinylation and pyridinylation would actually increase or decrease potency (along with expected improvement in accumulation).

Would really appreciate anyone's thoughts or suggestions on my analysis, and please correct me if I am wrong!

Many thanks,
Yuhang

[mattodd]

Tidying up old notes from the meeting:

1. To check concentrations of Yuhang's molecules in the biology assays, should we obtain extinction coefficients for them? @AJLloyd105
2. Following initial assessment, microbiology assessment is scheduled for 30-X-P, 85-2-P, 86-X-P and 88-X-P. Great.
3. Of the other compounds, Eve Carter's compound OSA1167 is looking interesting.
4. Was it the case that anything with a <20uM value can be looked at via crystal soaking.
5. OSA1145, 1164 and 1155 also looking interesting.
6. Can we do protein mass spec with any compound that looks like it might be a covalent binder?
7. Mention was made of using Diamond for time-resolved crystallography, for intermediates? Can someone expand on what and why? @LauraDS1
8. We need slides from the NEU team, ideally.