It’s how you move that matters – studying protein vibrations

giphy (1)

Molecular interactions resemble a dance, a composition of energy and motion.

A novel tool that may provide fresh perspectives to structural biologists has been developed by Dr Andrea Markelz’s group from the University of Buffalo. Its application towards understanding a biological function was published in  “Moving in the Right Direction: Protein Vibrational Steering Function” in the Biophysical Journal, with Dr Katherine Niessen as first author.

The group developed a technique called anisotropic terahertz microscopy (ATM) which was able to distinguish directional motions of protein vibrations. This is opposed to traditional methods that only measure total vibrational energy distribution by neutron inelastic scattering.

One way distal mutations (i.e. mutations far away from the site of disruption) affect ligand (i.e. drug/protein/other molecule) binding is by inducing long-range motions in protein structure that allows accommodation of the ligand. This technique could therefore provide a fresh look at drug-protein interactions, allowing scientists to discern and model drugs that not only bind but produce the desired vibrations to cause a particular effect.

Niessen et al. demonstrated the utility of ATM by studying the chicken egg white lysozyme (CEWL) and its binding to inhibitor (tri-N-acetyl-D-glucosamine, 3NAG). Binding of the inhibitor did not produce much change in the vibrational density of states (VDOS or energy distribution) whereas ATM could detect dramatically different changes between bound and unbound states.

They attributed these differences to the direction of vibrational movement. By computer modelling and simulation, the unbound (apo) form exhibited clamping motions around the binding site while the bound (holo) form displayed more twisting motions. Quite a stark difference when you consider the dance moves in “Heyyy Macarena” from “Twist and Shout”.

Furthermore, CEWL with distal mutations that induced higher catalytic activity without changes in binding, saw no significant change in VDOS but distinct differences in anisotropic absorbance. This demonstrates the utility of ATM to evaluate long-range mutations and their effect on protein activity.

Challenges remain as highlighted by Dr Jeremy Smith, a key expert in the field. This involves the need to crystallize and align proteins, and evaluating the detection sensitivity of the technique on a range of vibration types and under various conditions. But even he agreed it was a step (or shimmy) in the right direction.



Trump’s proposed budget

Trump released a budget blueprint last week that proposed rather drastic cuts in research and development, most of which would be channeled towards defense (in the form of wall building and boosting military defenses). Although it has to get through Congress later in the year to become reality, there is no doubt that the livelihood of scientists and the state of scientific research in America is coming under attack by Mr Trump. Here’s a brief summary of his proposed plans:

Trump’s proposed 2018 budget for R&D reflected in terms of Institute, Proposed budget change (% change from budget in 2016)

  1. National Institute of Health, NIH: $5.8 billion  (-20%)
  2. Department of Energy (Science), DOE: $900 million (-20%)
  3. Department of Energy (Advanced Research Projects Agency-Energy): $300 million (-100%)
  4. NASA Earth Science: $102 million (-5%)
  5. National Oceanic and Atmospheric Administration, Oceanic and Atmospheric Research (NOAA-OAR): $250 million (-50%)
  6. Environmental Protection Agency (EPA): $2.5 billion (-31%)
  7. Department of Energy (National Nuclear Security Administration): $1.4 billion (+11%)
  8. Homeland Security: $2.8 billion (+7%)


Matt Hourihon’s article nicely sums up which scientific programs would be affected together with more informative charts.

The basic goal of the budget was to increase defense capital by $58 billion (+10%). To do so without incurring significant debt meant reducing spending on non-defense capital by $54 billion, though that still leaves $4 billion unaccounted for.

Trump not only cuts funding for science but nearly every other area including housing and urban development, anti-poverty measures, agriculture, transportation and education. Read Vox’s article for an overview.

Although funding towards basic research and whether it drives economic growth has always been debated, the motivation behind the proposed steps seems driven more by xenophobia than by “making America great again”.

Trump seems to acknowledge that industry support for scientific endeavors are currently strong, which was his reason for completely de-funding grants for energy research. However the large cuts in basic science highlight his lack of understanding or even care towards the role a government plays in setting the climate for scientific research.

Ultimately, his actions would likely produce a brain drain in America. Thanks not only to the looming lack of research funding, but a general growing discomfort of foreign researchers feeling rather unwelcome.

I’m just glad to be living in a land where the leaders believe in the importance of science and share similar beliefs to Albert Einstein who said:

“Concern for man himself and his fate must always form the chief interest for all technical endeavours … in order that the creations of our mind shall be a blessing and not a curse for mankind.”


6 ways to keep up with scientific literature

So much information, so little time. Living in the digital age means information is easily available at the click of a mouse but it also means having to contend with a flood of news, views and reviews that can be overwhelming and confusing. Relevant scientific news these days not only comes in the form of journal publications but also pre-prints (see bioRxiv, pronounced bioarchives), Letters, blogs, news websites, and even God forbid, Tweets.

I’m ignoring (and secretly hating) the folks who have no problems keeping up, and whose answer to this question would probably just be to “read more”. For those like myself who have non-photographic memories and who tend to easily forget things previously read, I’ve put together a short list of pointers that would hopefully prove useful.

1. Use a citation software/web clipper

Mendeley is great for this, though I’m sure there are others. This allows you to easily download and sort your papers into folders, while enabling easy bibliographical citation. You can tag them with keywords, which adds another level of organization. A highly useful keyword search through the text of all the papers (provided you downloaded a pdf copy) is also available. It takes a while to get used to reading on a computer, but this really pays off in the long run as you don’t accumulate wads of space/tree-consuming paper that often end up unsorted and unread. A fullscreen mode on Mendeley allows you to read with a notes bar on the side that lets you highlight text and add comments.

For non-scientific articles, Evernote is great. Similar to Mendeley, you can sort web articles, photos, all kinds of media really, into folders and tag them with keywords. I love the clip from web function where a button at the top of your internet browser allows you to clip the website in simplified format. Often used when I’m surfing the net for ideas on what to blog about!

2. Create feeds/alerts

I spotted one of my Professors using HighWire for this and its a pretty nice one-stop shop for creating feeds/alerts. You can set up citation alerts that identifies journal articles containing your specified keywords/from specified authors and sends a list of their titles to your email. You can also sign up to receive an electronic table of contents from your favorite journals that allows for a more broader review of the recent literature. Recently, there have been some warnings that HighWire may be discontinued but so far these emails are still coming. I also am trying PubCrawler as a backup. eTOC alert emails can also be done directly at your favorite journal’s website.

Of course this only works if you spend the time to go through these alerts. Often what happens it is all these emails accumulate in your inbox collecting virtual dust. So best to set aside some time in a week to go through them and pick the articles of interest for more in-depth reading.

3. Get on Twitter 

Although I personally have not mastered the art of Tweeting, Twitter is an amazing resource for obtaining real-time insight into what key players in your field of study are talking and thinking about. Follow your scientific idols, and see who they follow, and follow them too. Not every scientist is on it though, but you’d be surprised sometimes at who you may find.

4. ResearchGate

I’m not a big user of ResearchGate, but they offer access to articles that you may otherwise have to pay for which is what drives many to get on it. Its a good way of seeing who has published what, who their closest collaborators are, and enables social interaction via online forums.

5. Write a blog or a literature review journal

Although reading widely is great for keeping up with literature, often it is remembering what you have read that is the challenge. A good way of cementing what you’ve read is by summarizing it and writing this down. It’s one of the reasons I started this blog. Often, I find myself searching and re-reading old posts to recall certain things. Writing a blog not only helps sort through your key thoughts, its a good way of collecting various sources of information into one easy-to-digest article, written in your own hand. If you’re shy about publishing it, create a private one. You’ll find yourself returning to it over and over.

6. Create a journal club

Interacting with people is naturally more memorable than reading something in private. Having a physical discussion about a paper in a coffee house or over food could help in remembering what was said, as odour memory seems to be the most resistant to forgetting. Furthermore, hearing opinions of your peers on the study also widens ones perspective. Even if the discussion is not physical, there are plenty of online forums, webchats and email threads that one can start with a group of people. In addition to the potential for generating new ideas, it’s a great way of keeping in touch!




What would you do with 900 million dollars of start-up funding?

America, a land of plenty – plenty of land, plenty of food, plenty of crazy politicians and plenty of start-up funding.

Grail, a company formed by sequencing giant Illumina in Jan 2016, recently obtained a hefty $900 million in Series B financing, after already obtaining $100 million in Series A. Grail aims to screen for cancer mutations in circulating tumour DNA (ctDNA) from blood samples via next-generation sequencing (learn more about this booming field in Sensitive Detection of ctDNA). The money came from several large pharmaceutical companies, Johnson & Johnson purportedly with the largest investment followed by others such as Bristol-Myers Squibb, Celgene and Merck. Interestingly, Bill Gates and Jeff Bezos from Amazon has also invested in Grail, together with the venture arm of medical distributor McKesson, China-based Tencent Holdings, and Varian Medical Systems, a radiation oncology treatment and software maker from Palo Alto.

This is the biggest start-up financing deal in biotech by a long-shot, the largest deal in 2016 went to Human Longevity, Craig Venter’s company that raised $220 million in series B. Another one that came somewhat close was RNA company Moderna Therapeutics, which raised $450 million in 2015.

Grail plans to carry out “high-intensity sequencing” on blood samples from vast numbers of people to detect circulating tumour DNA at early stages, essentially in people not showing any signs cancer, as a means of early detection to enable better treatment. This is an especially challenging feat, given that ctDNA makes up < 1% of circulating DNA found in the blood. But there are some hints that Grail is sitting on promising data sets that have turned skeptics into believers.

There are concerns that testing healthy people for cancer might yield false positives that could subject people to unnecessary and potentially dangerous testing procedures and treatments. This was the case for the Prostate-specific antigen (PSA) test used to screen men at risk for prostate cancer. It turned out that PSA testing did not significantly reduce mortality of men with prostrate cancer but did increase the harms associated with the accompanying treatments and tests, some of which are pretty nasty such as urinary incontinence and erectile dysfunction.

So Grail had better be sure the sensitivity and accuracy of their predictions are full-proof as cancer treatments are not exactly pleasant. They seem to be taking it seriously, judging from their embarkation on an ambitious trial called “The circulating cell-free genome atlas study” where they will recruit more than 10, 000 participants – 7000 newly-diagnosed cancer patients of multiple solid tumour types who have not undergone treatment, and 3000 healthy volunteers.  The trial is already recruiting and is projected to be completed within 5 years by Aug 2022, with a primary outcome measure available by Sept this year. Grail hopes to detect shifts in cancer stage severity as they perform their tests over time. How accurately their tests reflect other clinical readouts would give appropriate proof of its reliability. Likely, more trials involving more patients would be necessary to determine if this form of testing is full-proof and whether it may even replace tissue biopsies as a gold standard in cancer diagnosis.

Grail has even drafted plans to make their form of testing available to the medical community by 2019, subject to experimental results. An incredibly ambitious timeline, so its no wonder they need the big amounts of cash to drive it through. Jeff Huber, a former Google Exec, is Grail’s new CEO. His wife Laura died of colorectal cancer, so his new job also fulfils a personal mission. Other members of the team include other former employees from Illumina and Google, including Verily CSO/founder Vikram Bajaj. The Google Life Science company Verily have recently received a similarly outstanding investment of $800 million from who would have guessed, Singapore Temasek Holdings.

The scale of investment in America seriously dwarfs that found in the European biotech scene. Despite conservatives highlighting a potential bubble in US biotech and Trump’s anti-pharma sentiment that may signal a potential decline in available funding, one cannot deny that the lofty research goals being currently undertaken, can only yield an incredible expansion of scientific knowledge. In my opinion, science is expensive, and the more money you have, the more science you can do. They key thing though, is to make sure its good science!



An exciting time for epitranscriptomics

Epigenetics is a well-established means of gene regulatory control where chemical modifications of DNA bases or their associated histone proteins affect the expression of genes (as opposed to DNA sequence). These epigenetic marks may take the form of methylation (usually on cytosine residues) or histone acetylation/phosphorylation/ubiquitination/sumoylation etc. and can be passed down to daughter cells. Epigenetic changes are induced by the environment and provide a biological mechanism by which nurture as opposed to nature, plays a significant role in shaping our behaviours and characteristics.

Recently, scientists are uncovering the functional significance of epitranscriptomics – or the chemical modification of RNA. The first report of mRNA modification, specifically,  methylation of the N6 position of adenosine (m6A), occurred in 1974 by Fritz Rottman et al.. The m6A modification is the most common eukaryotic mRNA modification however its functional significance remained unclear until the 2010s. In 2012, Dominissini et al. using m6A-antibody enriched RNA-seq, discovered 12000 methylated sites in 7000 coding genes and 250 non-coding genes. Typically highly concentrated around stop codons, within long internal exons and at transcription start sites, it became evident that genes without these modifications tend to be more highly expressed. He writes about it in a Science essay here.

There are several players involved in epitranscriptomics, and they are often referred to as writers, readers and erasers. A methyltransferase METTL3 (“writer“) produces the m6A modification, the YT521-B homology (YTH) domain family of proteins are “readers” that bind to m6A modified RNA and regulate functions that affect protein expression such as RNA degradation, translation and splicing. Finally the “erasers” such as enzyme FTO, implicated in various diseases such as cancer and obesity, removed these m6A marks. Thereby completing the set of actors required for establishing epitranscriptomics as a regulatory mechanism for RNA expression.

Samie Jaffery’s group recently published a controversial paper that identified another epitranscriptomic modification N6,2O-dimethyladenosine (m6Am), located near the 5′ caps of mRNA, that is the main substrate of eraser FTO instead of m6A. m6Am is correlated with increased mRNA stability as it makes 5′ caps harder to remove, which thereby increases protein expression. This is in contrast with m6A modification, which is associated with suppression of protein expression. This highlights distinct functional roles of these RNA methylation marks.

There are already studies demonstrating m6A regulation is utilized also by non-coding RNA. The simple self-made schema below shows how m6A modification was used by Xist to carry out its transcriptional repression of the X chromosome, demonstrated by Patil et al. in a recent Nature publication.


Epitranscriptomics is a relatively young field and opens up new possibilities to study how RNAs are regulated and in particular, how non-coding RNAs may carry out their functions. This signals yet more exciting times ahead for RNA researchers!

Two man-made nucleotide bases produces never before seen semi-synthetic organism

As if altering the genetic code was not enough, now there might be a possibility to completely change the way it was written. Two new nucleotide bases have been created that were stably replicated in bacteria, expanding the possible number of base-pairs to 3 and increasing the coding capacity of DNA exponentially.

In his initial study published in Nature, Dr. Floyd E. Romesberg, a Chemistry Professor at the Scripps Research Institute in California, produced two nucleoside triphosphates called dNaMTP and d5SICSTP, which basepair via hydrophobic interactions instead of hydrogen bonding. A nucleotide transporter had to be expressed in the bacteria in order for their uptake, but once inside, DNA polymerases had no problem incorporating and amplifying the synthetic nucleotides, making copies of an introduced plasmid containing a single base-pair of the unnatural bases.

There were concerns initially that DNA repair mechanisms may work to remove these foreign bases, but that did not seem to be the case. The unnatural bases persisted for more than 3 days after he stopped supplying the bacteria with nucleotides at a retention of 45% (at 0 days, retention was in excess of 95% as suggested by termination of reads by Sanger sequencing) and 6 days at 15%. This led him to guess that their decay over time was due to replication-mediated mispriming instead of DNA repair.

In his most recent study in PNAS, Dr. Romesberg and his graduate student Yorke Zhang made improvements to the nucleotide transporter used, which previously induced some toxicity. They also further optimized one of the unnatural bases to be better incorporated by DNA polymerases, increasing copying efficiency. Finally, to prevent the decay of the unnatural bases over time, they used CRISPR to eliminate the DNA that had lost the unnatural bases. The resulting bacteria was able to indefinitely retain the unnatural bases under various sequence context.


Image taken from News Medical Life Sciences‘ interview with Professor Romesberg

The so-called semi-synthetic organism produced therefore uses 3 base-pairs and has 6 nucleotide types instead of 4. It remains to be seen whether its DNA can be transcribed into RNA and then more importantly into protein. But it holds the potential to greatly change the way things are currently done – imagine what we could do with 172 amino acids instead of 20!


Role models in science – Dr Susan Lindquist


It has been a month since renown researcher of protein folding, Dr Susan Lindquist, passed away to cancer at the age of 67. I remember watching her video on prion biology while working on my PhD project looking at mechanisms involved in neurodegenerative disease. She seemed pretty cool, I thought.

She grew up in Chicago to a Swedish father and Italian mother who never expected her to come so far in her career. Being a woman, their hopes for her was to marry someone decent and successful. Coming home from a party the night before New Year’s Eve and seeing their daughter hard at work on a paper, their comments were “Are you still working? When are you going to settle down?”  I’m not so sure that parental thinking has changed significantly since then…

She found inspiration in a book detailing the life of Elizabeth Blackwell, the first woman who obtained a medical degree in the US, and various teachers who stimulated her interest in science. Under the guidance of her microbiology professor Jan Drake, she applied successfully for a National Science Foundation scholarship to do research in his lab. With his encouragement, she applied to graduate school in Harvard, and got in, something she had never dreamed would happen. She worked in the lab of Matthew Meselsen but failed to get any data for her first project. After talking to a colleague down the hall who noted particular phenomena to heat-exposed fruitflies however, she decided to test if any similar responses would happen in cells. That was the turning point in her career, as she found and characterized the upregulation of specific proteins induced by heat, a mechanism termed the heat shock response that would be found to be highly conserved across many organisms.

She continued to work on the heat shock response during her post-doc rather independently in the lab of Hewson Smith at the University of Chicago. She characterized how the expression of these heat shock proteins were regulated via transcription, translation, splicing or degradation. Realising that these proteins were so highly conserved across different species and were being in expressed in every cell in response to stress that occurs frequently in life and disease, Lindquist was driven to find out exactly what these proteins were doing. Her research brought her into broad and vastly different fields, as it was found that these proteins played essential roles from enhancing malignancy in cancer to managing protein aggregates so often found in neurodegenerative diseases.

She had surpassed her initial dream of writing grants under the supervision of a male superior, to managing her own lab at the Whitehead Institute at MIT. She even co-founded a company – FoldRx Pharmaceuticals – which utilized her favourite model, yeast, in a high-throughput functional assay to search for drugs that could alleviate protein aggregation in protein misfolding diseases. This was later bought by Pfizer as they sought to obtain the rights to the drug Tamafidis, which was approved for the treatment of early stage transthyretin-related hereditary amyloidosis or familial amyloid polyneuropathy or FAP.

Susan Lindquist is definitely a role model to look up to, especially for women in science. There are still far lesser women compared to men in leadership positions in science and beyond. I gather this is attributable to the demands of family rearing, the discrimination that comes hand-in-hand with being a woman attempting to lead, and the internal fight women go through to overcome natural feelings of inadequacy. But Susan shows us it can be done. And I think we would probably do a better job than men sometimes as Sandi Toksvig would agree in her hilarious TED Talk.

Read and watch more about Dr Susan Lindquist here:

Fearless about Folding | The Scientist Magazine®

Gitschier, Jane. “A Flurry of Folding Problems: An Interview with Susan Lindquist”. PLoS Genetics. 7 (5): e1002076. doi:10.1371/journal.pgen.1002076. PMC 3093363Freely accessible. PMID 21589898.

Short video Q&A with Susan Lindquist