Codon frequency optimization demonstrates the anti-virus capability of SARS-CoV-2

A study conducted by researchers at Ohio State University and New York University’s Grossman School of Medicine in the U.S., has revealed that codon optimization for bacterial sensitivity causes the production of a non-structural enzyme inactive protein 12 (Nsp12), a catalytic subunit of RNA polymerase RNA-dependent respiratory distress syndrome coronavirus 2 (SARS-CoV-2). The study is currently available on the bioRxiv* preprint server.

Background

SARS-CoV-2, the causative pathogen of coronavirus infection 2019 (COVID-19), is a single-stranded, positive RNA virus with a genome size of about 30 kb. In addition to encoding immunogenic structural proteins such as spike, membrane, nucleocapsid, and envelope proteins, the SARS-CoV-2 genome encodes several nonstructural proteins (Nsps) that are essential for viral reproduction and expression gine. In as active a state as possible, the RNA polymerase RNA (RdRp) of SARS-CoV-2 consists of Nsp12, Nsp7, and Nsp8.

In the present study, the scientists performed a structural and functional analysis of SARS-CoV-2 RdRp.

Important comments

The scientists used the E. Coli expression platform to purify essential proteins and evaluate the mechanical details of RdRp. They observed that RdRp collected by purified Nsp12 remains inactive. On further analysis, they identified that Nsp12 recall was the main reason for this low level of RdRp activity.

While examining the cause of low activity level, they found that the enzymatic activity of Nsp7-Nsp8-Nsp12 is still very low on various templates, such as the optimal hairpin scaffold. After a detailed examination of the cleaning process and the reaction conditions, they removed the His₁₀ tag attached to Nsp12 for purging. However, His removal₁₀ tag did not increase the activity of the enzyme.

Furthermore, they assumed that the decrease in the level of RdRp activity could occur as a result of recurrent protein misfolding. Any change in the coding mRNA could be a possible cause of low enzymatic activity. When Nsp12 is made using the E. Coli sensing platform, the codon sequence of Nsp12 is usually modified to conform to the host codon usage. It is common practice despite the fact that even codon substitution can dramatically alter protein activity.

To test the hypothesis, they performed a series of experiments and found that RdRp collected with active Nsp12 has a much higher activity level than the level collected by purified Nsp12. In terms of protein misalignment, they observed a large structural difference between active and purified Nsp12 in terms of access to several domains of RdRp.

The team’s comparative analysis has revealed that there are two regions with rare codon accumulations in the active Nsp12, but not in purified Nsp12. Rare codons are essential to stop the ribosome and to ensure that proteins fold properly. By constructing chimney proteins by substituting corresponding segments of purified Nsp12 for those segments of Nsp12, they found that chimney proteins with codon 350-435 are derived from inactive purified Nsp12.

These observations indicate that proper translation of the 350-435 region is essential for optimal Nsp12 activity and Nsp7-Nsp12 interaction, which in itself is essential for the proper functioning of RdRp.

With further structural analysis, the scientists noticed a difference between active and purified Nsp12 in the NiRAN land layer, which is essential for nucleotide binding. The scientists have therefore suggested that this variation in protein folding may be due to the difference in active and purified Nsp12 functions. Using E. Coli strain with mutated ribosomal protein S12, they have found that proper translational compression of Nsp12 can be possible.

Investigate meaning

The study reveals that codon frequency (retrieval) optimization of Nsp12 mRNA to support bacterial protein expression can lead to protein malformation, which may significantly reduce SARS-CoV-2 activity RdRp. The presence of rare codons in mRNAs is essential for protein folding.

In other words, the study suggests that translation of undeveloped mRNA can generate highly active RdRp, and these findings are particularly valuable for functional studies, for the development of sensory platforms. bacterial appropriate, and more importantly, for the identification of novel RdRp inhibitors.

* Important message

bioRxiv publish preliminary scientific reports that are not peer-reviewed and, therefore, should not be seen as final, guiding health-related clinical practice / behavior, or be treated as information established.