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Archive for the ‘Glycobiology’ Category


Some months before @robField’s tweet setting off the train that led to the Eu sensor that discriminates PAP/PAPS, Ed Yates and myself were having a curry with Dulce Papy-Garcia from UPEC, who had examined one of our PhD students. A matter we discussed at length was ‘why sulfate’. That is, why does biology use both sulfate and phosphate to modify post synthesis proteins, polysaccharides and other molecules. We didn’t come up with an answer,  but the conversation led Ed and myself to consider that the question merited exploration. 

This we thought would be a simple matter. 

It turned out to be one of the most difficult papers Ed and myself have written, to the extent that after N drafts (where N is a significantly larger number than either of us had experienced in any previous writing exercise) and too many summers we still had nothing satisfactory. So, we cunningly inveigled two colleagues, Tim Rudd from NIBSC and Marcelo Lima from Keele to join us on what we advertised as the sunny beach of sulfate and phosphate, but which in reality was a rather dank quagmire. There is though something about strength in numbers, and with very helpful input from Steve Butler in Loughborough, we arrived at what we considered a satisfactory synthesis. Happily, the reviewers concurred, and the paper is now published at Royal Society Interfaces, “Phosphorylation and sulfation share a common biosynthetic pathway, but extend biochemical and evolutionary diversity of biological macromolecules in distinct ways”.

This is by no means the last word on the matter, but along with some previous thoughtful papers we cite (if we have missed one, please let me know) it provides some ideas that may help us to understand why biology co-opted particular elements from the inorganic world to perform groups of functions vital to life as we know it now.

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More sulfation

Earlier this year Simon Wheeler (who now has a well deserved substantive position, congratulations!) and Steve Butler published the first output from the BBSRC TDRI awarded to Steve, with myself and Ed Yates in supporting roles. It is always nice to collaborate with real chemists, as it reminds me I am very much a pseudo chemist, and I learn a lot. After what I would consider a quite heroic effort on the synthesis front, Simon and Steve pulled out a very useful sensor, based on a europium complex. The Eu sensor has good selectivity for PAP over PAPS, the universal sulfate donor. The assay works well and is very amenable to high throughput 384 well format assays (= more papers on the way). So we can now measure sulfotransferase activity in realt-ime independently of the acceptor for pretty much any enzyme-substrate combination. This represents an important tool for the wider sulfotransferase community. 

The paper also demonstrates the importance of social media in science, as a means to access in a non-direct manner new information that sets off an innovative project. I saw tweet from @Fieldlab highlighting a paper from Steve’s lab on lanthanide sensors able to discriminate nucleotide phosphates and read the paper. Naively I thought PAP/PAPS sensing using such compounds should be easy, so I contacted Steve. After some preliminary tests with PAP and PAPS on his side, we wrote the grant – another lesson here, as the application neared final from I went over to Loughborough for a meeting, which allowed us to iron out a few problems far more effectively than by electronic communication. The work was, as hinted above, far from straightforward, but like everything that is new, very rewarding and continues to be so.

I have just moved from the bird site to the proboscidean one and things look like there will be even more of such ‘random access’ of information there, so let’s see what turns up!

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After an interaction on Twitter with a colleague who is associated with #LongCovid and #TeamClots, he asked me for some references. I thought what to send, and then realised that references plus something a bit more than a Tweet might be useful, so here goes.

(more…)

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With new data in hand, our first preprint on SARS-cov-2 receptor binding domain (RBD) interacting with heparin now has a sibling, which demonstrates that heparin inhibits the infection of Vero cells by SARS-cov-2

Some of the key points of the team’s new work are:

  1. Inhibition of viral infectivity in a Vero cell model by heparin, which is a better inhibitor for SARS-cov-2 than SARS-cov.
  2. Analysis of the interactions of a more extended library of model heparins with the SARS-cov-2 receptor binding domain. As with many other heparin-binding proteins, these data show that while sulfation is critical for RBD binding, the amount of sulfate is not, but instead it is the spatial arrangement of sulfate groups that is most important.

Together the data point to heparin being a potentially useful therapeutic to reduce infectivity. (more…)

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E-mail I sent out to the research group today.

Dear All,

From now on we really do need to reduce lab work and enforce strictly social distancing, something we stated last week.

The first transmissions from Scousers who picked up the virus on match night (bars, clubs, hotels) from Athletico fans will occur this coming week and next week; we will then get F2, F3 (F number related to contact: primary = F1, a contact’s contact = F2 etc.) transmission. (more…)

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Thursday last week (Feb 27) Mark was up from Keele and popped his head around my office door – not a surprise, as he is often here to do circular dichroism on various heparin-binding proteins – to announce that Marcelo had managed to make some SARS-CoV-2 S1 receptor binding domain. Mark had asked Hao,  my postdoc, to do some SPR measurements to see if it bound heparin.

Later in the day I went over to the SPR/CD lab to find Courtney, Mark’s PhD student and Mark beavering away on the CD. A quick discussion. Hao had finished some work on our first grade A heparin functionalised SPR surface, so we set about injecting the SARS-CoV-2 surface protein (Spike) S1 Receptor Binding Domain – a one shot experiment, as amounts of protein were limited, so we injected 1 mL at 500 µL/min (I like high flow rates as mixing is way better, though still far from perfect).

Bingo. (more…)

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I have written the following to my Brexit Party Representatives in the European Parliament. At the heart of parliamentary democracy is the idea that our representatives do indeed represent the interests of their constituents, regardless of Party politics. Of course interests have to be balanced but when these are win-win, there can be no reason for not representing a particular interest. As the matter is not a personal one, then I have also put it on my blog, since there is no reason for secrecy.

I will, of course, post any further correspondence, unless it is confidential for some reason.

“Dear Ms Fox (& cc’ed to Mr Nielsen and Mr Bull)

I am writing to you as one of your constituents regarding an issue which affects me personally and the region. As a University Professor I have over the years been awarded research funds from various Framework Programmes. Most recently, I am part of the €4M FET-OPEN programme “ArrestAD”. This aims to test a new paradigm for Alzheimer’s Disease screening and lay the foundation for a new class of drugs that would arrest the disease.

FET-OPEN projects are very much blue skies and in our case we appear to have hit the jackpot. The trials of the diagnostic in Paris and Warsaw  are quite spectacular and our own work has shown that the targets which ArrestAD has proposed are eminently druggable.

ArrestAD is a 4 year programme. As in any blue skies research, towards the end of the penultimate year a decision is made by the research team whether we should apply for a new, larger project, under one of the translational programmes available under H2020, or to can the idea, in the event it isn’t going to deliver.

Since ArrestAD is delivering its promise, the team will be going forward and applying for a translational programme. This will involve further clinical centres and greater industry participation, since we need more patients and, for drug development, far greater resource.

Alzheimer’s being what it is, ArrestAD obviously impacts widely and not just on myself: there are substantial social and economic ramifications for our region, the UK, and beyond.

The problem we face is that with a so-called No Deal Brexit, the UK loses access to funding from H2020 and the future framework programme. There is no  legislation in the UK Parliament that would guarantee funding as a 3rdcountry. The upshot is that Brexit will prevent my continued contribution to this likely life-changing research programme. Importantly, it will prevent the UK from reaping economic benefit (clinical trials, pharmaceutical industry).

As my representative in the European Parliament, it is imperative that you work to find a solution, which ensures that the drug development arm of this project remains based in the UK, and that the UK is able to participate fully in the wider clinical trials of the diagnostic. I think you would agree that given the impact of this dreadful disease, this is in everyone’s interests.”

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Friday Pat Eyers pushed our two papers on new screens we have developed for sulfotransferases up onto Biorxiv. More about the history of this work later. For now the briefest of summaries.

The heparan sulfate 2-O sulfotransferase paper is here

and the tyrosine sulfotransferase paper is here.

The key messages are:

(1) Mimetics of PAPS, the universal sulfate donor, that inhibit sulfotransferases are present in kinase inhibitor libraries.

(2) We demonstrate selectivity, in that some compounds inhibitor one sulfotransferase better than they do the other.

(3) PAPS mimetics look like providing a rich vein of sulfotransferase inhibitors of varying selectivity, rather like ATP mimetics have done for kinases.

(4) We have two very effective high throughput screens, which means no sulfotransferase is now beyond our reach.

Sulfation has been frustrating due to the lack of chemical tools to selectively inhibit a particular sulfotransferase. With these two papers we can foresee such tools in the not too distant future and with these, we can unpick the role of sulfation in biology, from development, through homeostasis to disease.

Exciting times!

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Congratulations to Aiseta!

On Monday 4 December Aiseta Baradji successfully defended her thesis. A long journey and a hard one as ever with its ups and downs, surprises and a certain amount of head scratching over data that push us in new directions. In the end a great thesis that will be consulted in the labs of her supervisors for a long time. Now onto the next phase.

 

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Two postdoc positions are available in my lab.

Both are part of the larger, European Commission-funded FET-Open programme, ArrestAD, which has recently been funded.

Position 1 aims to characterise heparin-binding proteins in Alzhiemer’s disease.

Position 2 aims to develop inhibitors to Golgi sulfotransferases. (more…)

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