Thread by @DrEricDing

‘ONE NEAT TRICK’ for #COVID19 vaccine:

Yes, sounds clickbait-y, but there actually was a special technique developed to enhance the vaccine to be more effective—by adding a “double proline” (2-P) to stabilize the spike.

Took years to discover it! Neat.
cen.acs.org/pharmaceutical…

2) “Graham’s lab had a template to work from because they’d spent several years designing an experimental mRNA vaccine for MERS.
But there’s a third, more subtle secret to their success: a tiny but oh-so-important tweak to a critical viral component called the spike protein.”

3) “Viruses multiply by dumping their genes into our cells and hijacking our cellular machinery to crank out new virus particles. But first, they need a doorway into our cells. Coronaviruses are studded with spikes, which grab hold of proteins decorating our cells like doorknobs.

4) “Once attached, the spike undergoes a dramatic transformation, stretching before partially turning inside out to forcefully fuse with our cells.
Scientists believe that for COVID-19 vaccines to be effective, our immune systems must develop antibodies that prevent this fusion.”

5) “Such antibodies must target the spike protein in its aptly named prefusion conformation. Unfortunately for vaccine developers, spike proteins are liable to spring from their stubby prefusion shape into their elongated postfusion form on a hair trigger.

6) “Fortuitously, Graham and a former postdoc, Jason McLellan, devised a solution to this problem before the pandemic. Through a bit of structural biology and persistent protein engineering, McLellan discovered that adding two prolines—the most rigid of the 20 amino acids—

7) “to a key joint of a vaccine’s spike protein could stabilize the structure’s prefusion shape. This 2P mutation worked in preclinical studies of Graham and Moderna’s MERS vaccine, so they applied it to Moderna’s COVID-19 vaccine.”

8) “As Norbert Pardi, an mRNA vaccine scientist at Univ of Penn, puts it, we’re “very lucky, actually,” that scientists worked out the 2P mutation for a MERS vaccine before the COVID-19 pandemic. “It wouldn’t be possible to go so fast with the Moderna vaccine otherwise.” 🙏

9) which other COVID vaccines use the 2P trick?

Johnson & Johnson, Novavax, and Pfizer. But not AstraZeneca (I don’t know why).

“The 2P mutation might quite literally be the smallest detail that could make or break the first generation of COVID-19 vaccines.”

10) The reason I share the article is to explain that the success of the vaccines was not an overnight thing. ➡️It took YEARS AND YEARS of testing to develop the perfect vaccine. There are many other laborious vaccine design details in each COVID vaccines like this. Be grateful.

11) MORE DETAILS OF THE 2P SAGA:

“Despite many attempts, and even with outside help, the scientists couldn’t get the MERS spike protein to cooperate. So McLellan, Graham, and their collaborator at Scripps Research, Andrew Ward, turned to a different coronavirus, HKU1...”

12) “The virus, which causes the common cold, was safer to work with and better at sitting still while its picture was being taken. In 2016, Ward’s lab used a technique called cryogenic electron microscopy to capture the structure of the HKU1 spike.”

13) “McLellan and his team scoured the structure for clues that could help them stabilize the MERS spike protein. They homed in on a region toward the top of the spike, a small loop of amino acids that held two coil-like structures called α-helices together.

14) ““It is like a spring bent in half,” When the spike protein binds to a human cell, that spring is released, and the two helices and the loop straighten into one long helix that harpoons the human cell and pulls the virus and human membranes close together until they fuse.

15) “Blocking the release of that spring should prevent viral fusion, giving the immune system a chance to make antibodies that prevent infection. McLellan’s lab tried several tricks before arriving at one that worked: adding two prolines in the loop clamps the spring together.

16) “This 2P mutation enabled McLellan and Ward to solve the MERS prefusion spike structure in 2017 (Proc. Natl. Acad. Sci. U.S.A., DOI: 10.1073/pnas.1707304114). Graham began working with Moderna to make an mRNA vaccine for MERS using the 2P mutation that same year.

17) “Once the genetic sequence of SARS-CoV-2 was released, Graham’s & McLellan’s lab were able to compare its genome with SARS and MERS, pinpoint the code corresponding to that bent spring, and then add the 2P mutation to lock the SARS-CoV-2 spike in its prefusion conformation.”

18) “After rapidly determining the structure of the SARS-CoV-2 prefusion stabilized spike protein in February, McLellan’s lab continued to tinker with the spike protein. After testing more than 100 variations, he published a new version in July called HexaPro.

19) “It contains the original 2P mutation plus four additional prolines that further stabilize other regions of the protein. His preliminary results suggests the molecule might be 10 times as potent as 2P.

20) ““There is probably no limit to how much more we can keep going or how much time we can keep spending,” McLellan says, but he’s happy with HexaPro. Multiple vaccine companies are already conducting lab tests of HexaPro as part of a second-generation COVID-19 vaccine.” 🙏

21) Here is what adding 2P does—its stabilizes the original spike protein, and prevents it from shape shifting to another shape that allows entry into human cells. Shapeshifting also makes antibodies more troublesome—so stopping the shape-shift is key! That’s what adding 2P does.