Hunt for COVID-19 ‘Heart Stopper’

SARS-CoV-2 (2019-nCoV) coronavirus main protease, with inhibitor in turquoise.

Know the COVID-19 Anti Viral development path to avoid fake news.

In this world of fast and easy travel, emerging viruses are increasingly becoming a major danger to world health. Coronaviruses are a notable example. Particularly virulent forms have emerged from their natural animal hosts and pose a threat to human communities. In 2003, the SARS virus emerged in China from bat populations, moving to civets and finally to humans. Ten years later, the MERS virus also emerged from bats, transferring in the Middle East to dromedary camels and then to humans. Recently, another coronavirus has emerged in China by way of animals in a live market. Structural biology is helping us understand these dangerous foes, and hopefully will help us develop new ways to fight them.

At the frontlines of this battle against COVID-19 are healthcare workers, who risk their lives daily as they screen, diagnose and treat those infected. But hard at work behind the scenes are our scientists racing to understand the virus, in hopes of finally ending the outbreak.

Structural biologists, in particular, play a crucial role. Like assembling the pieces of a particularly challenging puzzle, structural biologists use advanced imaging techniques to determine the intricate, three-dimensional structures of biological molecules like COVID-19, as seen above. Researchers can then use this knowledge to design or guide their search for novel antiviral drugs.

Coronavirus Proteases — A tale of two subunits

COVID-19 belongs to a subset of viruses known as coronaviruses — so called for the ‘crown’ of proteins that dot the viral surface. Underneath its royal exterior lies a lengthy strand of ribonucleic acid (RNA), which serves as the viruses’ genetic material.

When COVID-19 infects a cell, it hijacks the existing molecular machinery to create long chains of proteins required by the virus to generate even more copies of itself. These long viral proteins, however, only become functional when cut into smaller pieces by proteases.

Thus, coronavirus proteases like that of COVID-19’s play an integral role in propagating the virus. As seen in the diagram, its distinctive heart shape is the result of two identical protein subunits (colored orange and red) coming together to form a functional protease.

Similar to a lock and its key, the protease’s activity is triggered by the binding of molecules to specific points on the protease called active sites. The binding of a substrate effectively switches the protease on, allowing it to cut the long viral protein strands into smaller chains.

Searching for a heart-stopper in the same way, the protease’s activity can also be blocked by molecules called inhibitors. When an inhibitor (depicted in turquoise) attaches to an active site, it prevents the binding of substrates — stopping the action of the protease altogether.

Therefore, finding an inhibitor for COVID-19’s heart-shaped protease is the first step to beating the epidemic which is what the current research and trials being conducted is all about.

Alas, some things are easier said than done. Coronaviruses and their proteases are a diverse bunch, so inhibitors for one type of coronavirus may not work for COVID-19. But examining the protease structure of related coronaviruses, like those found in bats is helping identify inhibitors against COVID-19 and other emerging viruses. Another approach is to repurpose existing antiviral drugs.

For instance, a cocktail that includes HIV drugs lopinavir and ritonavir — both protease inhibitors — has seen early success in treating patients with severe cases of COVID-19.

Also Camostat Mesylate (+E-64d), the drug shown by German scientists to effectively block corona virus CoVID-19 infection of lung cells, is apparently made by Sun Pharma in India and sold as Camopan. It is approved for human use in Japan but is not FDA-approved.A significant concern though is that TMPRSS2 might not be the only protease that controls spike priming and hence blocking it may be ineffective in people as other proteases may act as backups, still allowing the virus entry into cells. TMPRSS2 is being repurposed. Its not a cure in itself especially due to the fact that this virus is a RNA virus that does not have its DNA. Which protein it shall grow using human DNA is huge open question for Camostat to be called a vaccine.

Coronavirus proteases are attractive targets for the design of antiviral drugs. Finding a way to block COVID-19’s distinctive heart-shaped protease is the key to stopping the ongoing epidemic.

Coronavirus Code

Coronaviruses contain a genome composed of a long RNA strand — one of the largest of all RNA viruses. This genome acts just like a messenger RNA when it infects a cell, and directs the synthesis of two long polyproteins that include the machinery that the virus needs to replicate new viruses. These proteins include a replication/transcription complex that makes more RNA, several structural proteins that construct new virions, and two proteases. The proteases play essential roles in cutting the polyproteins into all of these functional pieces.

Main Protease

The main protease of coronavirus makes most of these cuts. The one shown here is from the SARS-CoV-2 (2019-nCoV) coronavirus that is currently posing dangers in Wuhan. It is a dimer of two identical subunits that together form two active sites. The protein fold is similar to serine proteases like trypsin, but a cysteine amino acid and a nearby histidine perform the protein-cutting reaction and an extra domain stabilizes the dimer. This structure has a peptide-like inhibitor bound in the active site.

SARS Proteases

The two proteases from SARS are shown here. The main protease is similar to the Wuhan one, and cleaves at 11 sites in the polyproteins. The papain-like protease has single subunit and also uses a cysteine in the reaction. It makes three specific cuts in the SARS polyproteins, and also clips several proteins in the infected cell, including removing ubiquitin from ubiquitinated proteins.

One of the consequences of this deubiquitination is that it interferes with production of interferons in the innate immune system, short-circuiting some of our defenses against the virus.

SARS main protease (top) and papain-like protease (bottom), with inhibitor in turquoise.

Bat Coronavirus Main Protease with Inhibitor

Researchers are actively using these structures to search for compounds that block the action of the proteases, for use as antiviral drugs.

The diversity of coronaviruses poses a great challenge with this effort: coronaviruses have been classified into four separate genera, and sequence and structural studies have shown that the proteases of these viruses can be very different, so drugs designed to fight one may not be effective against others.

The active site cysteine and histidine are shown in the illustration, with an inhibitor in turquoise.

One possible way to address this challenge is to try to design a broad-spectrum inhibitor targeted against the progenitor bat coronavirus, such as the one shown here, which may then provide a head-start for discovering inhibitors against newly emerging viruses.

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