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Posts Tagged ‘heparan sulfate’


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.

From a bioinformatics standpoint, we would expect heparin to inhibit virus infection, since the ACE2 binding site on the protein overlaps the heparin binding site. Moreover, heparin was established to inhibit SARS-cov infectivity and the two viruses have good sequence homology in their RBDs, including the ACE2 binding site and the putative overlapping heparin binding site.

However, to determine if this is actually the case takes time. It is nearly a month since the the team’s initial preprint (now updated with low molecular weight heparins), which was the first demonstration of an interaction between the SARS-cov-2 receptor binding domain and heparin. In the ensuing month, the usual lab gremlins have been working overtime to thwart us. In particular we should cite the Scale up Protein Production gremlin, who is a really awkward so and so. Then there is the Devil’s very own side kick, the Surface Gremlin, who has the uncanny ability to mess up the most careful of preparations of sensing surfaces. Anyway, we know them well and wrestled them into submission, though it did take a couple of all nighters to finally send them packing to some other lab (rest assured, they will not stay with you forever, they know where we work and have 24/7 access to our building, they will be back here and leave you alone for a while).

We have a large library of sugars and sugar-related structures to investigate, courtesy of friends and colleagues in the glycoworld. Some relate to potential therapeutics, others to mechanism. So our work is far from finished on this front.

It has been a lot of fun, with the team’s work varying in intensity from the equivalent of lazing in the sun in front of the trench, to long stints in the lab, including a couple of 22 h ones.

It has also been profoundly rewarding. There are some strange features of the interaction of the SARS-cov-2 receptor binding domain with heparin, not the least of which is the need for SDS to regenerate heparin surfaces with bound SARS-cov-2 receptor binding domain, rather than the more usual 2 M NaCl. This is not unprecedented, as many moons ago one of John Gallagher’s team saw something similar with the interaction of thrombospondin with heparin – I think in that case we had to use 1 M urea to regenerate surfaces.

Cellular heparan sulfate is integral to the infectivity of many pathogens and exogenous heparins have been found to be effective inhibitors. In the case of respiratory viruses this extends beyond the coronaviruses to include influenza viruses. HIV and herpes have long been known to be inhibited by heparins, while more recently this has been shown for Zika and other flaviviruses. On the parasite front, Plasmodium falciparun rosetting is inhibited by heparin and a literature search using your favourite pathogen and ‘heparin OR heparan sulfate’ will pull out many more examples.

Somewhat surprisingly, particularly since heparins lacking anticoagulant activity show good inhibition of pathogens, none of this has hit the clinic yet. There are a number of reasons behind this. Heparin and its derivatives are not single chemical entities, which makes companies nervous in relation to regulatory approval. Moreover, this is not blockbuster territory, partly because once one heparin drug is in use, it will not be so difficult to make another that is not covered by the patent. This relates in part to the fact that heparins are not single chemical entities and the myriad routes to making derivatives. There are, for example, a number of different proprietary low molecular weight heparins, though these are not clinically equivalent. Then there is the market. Many of those in need of such therapeutics are poor and a ‘blockbuster’ drug with a hefty price tag is not going to have much of a market.

On the plus side, it is unlikely that a heparan sulfate-dependent pathogen will be able to evolve resistance to a heparin therapeutic – heparan sulfate is too fundamental to metazoan biology for it to be bypassed. So perhaps covid19 will mark the point at which heparins start to be tested clinically as therapeutics in infectious diseases.

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Today was the culmination of Zaid’s PhD journey, when he successfully defended his thesis on the use of gold nanoparticles to probe the mechanism of action of a peptide that inhibits ‘flu virus infectivity. Though he approached his viva with trepidation, his beaming face afterwards told a different story. The usual smattering of corrections, a paper already up on Bioarxiv ready to submit for peer review and another to put up on Bioarxiv, and in a few weeks he will be truly done, with a CV to match.

 

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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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Our review on fibroblast growth factors (FGFs) as tissue repair and regeneration factors, which we made available as a preprint from the time of submission is now published at PeerJ. (more…)

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Of nanoparticles, cells and polyanions

It is the end of semester 2 so it’s marking season. Since we double mark (a good thing), the final year research projects are marked by both supervisor and an assessor, a member of staff who is not involved in the project. One of the projects I marked was Gemma Carolan’s on “How do SmartFlares RNA detection probes reach the cytosol? Available are the PDF of report, and posts here and here.

I had a sense of déjà vu while reading the project – the clear endosomal location of the SmartFlares, regardless of the DNA sequences brought me back to the days when antisense was the technology of the future for medicine.

While evaluating new technology it is useful to go back and look at other high flying technology. The reality is that it takes decades before we know whether the promise (and hype) were justified; this is true for any hot topic from stem cells to nanoparticles and graphene.

Antisense effects can be mediated by RNAse H, an enzyme that specifically cleaves RNA-DNA duplexes and which protects our cells from RNA viruses. There are other mechanisms, e.g., interference with splicing or translation, but the RNAse-H mediated transcript degradation should be central to many antisense effects. There were many papers reporting specific effects (evidenced by differences between sense, antisense and scrambled oligonucleotides sequences). These certainly contributed to success of individuals and of institutions, e.g., in UK Research Assessment Exercise and grant awards.
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I am a fan of PubPeer, as it provides a forum for discussion between authors and the wider community, something I have discussed in a number of posts (two examples being here and here). Two days ago, My colleague Mike Cross came by my office, having just delivered a pile of exam scripts for second marking (it’s exam and marking season), asking if I had seen a comment on our paper on PubPeer. I had not – too many e-mails and too busy to look at incoming!
So I looked at the question, which relates to panels in two figures being identical in our paper on neuropilin-1 and vascular endothelial growth factor A (VEGFA) – indeed they are labelled as being identical.
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