Molecular Interactions of Zyesami with the SARS-CoV-2 nsp10/nsp16 Protein Complex.

BACKGROUND: SARS-CoV-2 emerged in late 2019 and caused COVID-19. Patients treated with Zyesami were found to have a 3-fold decrease in respiratory failure and improved clinical outcomes. It was reported that Zyesami inhibits RNA replication of SARS-CoV-2, including several non-structural proteins essential in viral RNA replication. SARS-CoV-2 is a distinctive virus that requires nsp10 and nsp16 for its methyltransferases activity which is crucial for RNA stability and protein synthesis. OBJECTIVE: We aimed the in silico determination of inhibitory consequences of Zyesami on the SARS-CoV-2 nsp10/nsp16 complex. Targeting SARS-CoV-2 nsp10/ nsp16 protein complex may be used to develop a drug against COVID-19. METHODS: I-TASSER was used for secondary structure prediction of Zyesami. CABS-dock was used to model Zyesami with SARS-CoV-2 nsp16 interaction. The docked complex was visualized using PyMol. The quality of the docking model was checked by using ProQdock. RESULTS: The 3D structure of SARS-CoV 2, nsp10/nsp16 showed that essential interactions exist between nsp10 and nsp16. Significant contact areas of Zyesami exist across amino acid residues of nsp10; Asn40-Thr47, Val57-Pro59, Gly69-Ser72, Cys77-Pro84, Lys93-Tyr96. In addition, polar contacts between nsp16 and Zyesami are Asn299-Ser440, Val297-Asn443, Gly149-Tyr437, Gln159-Lys430, Asn178- Arg429, Ser146-Arg429, Ser146-Arg429, Lys147-Arg429, Asr221-Thr422, Lys183-Asp423, Lys183-Asp423, and Gln219-Asp423 the residues are shown of nsp16 and Zyesami respectively. CONCLUSION: The structural bioinformatics analyses have indicated the potential binding specificity of Zyesami and nsp16. Data predict how the initial binding of Zyesami with nsp10 and nsp16 may occur. Moreover, this binding could significantly inhibit the 2 -O-MTase activity of the SARSCoV nsp10/16 complex.

Low pH up-regulates interleukin-6 mRNA in L6-G8C5 rat skeletal muscle cells independent of pH sensing by SNAT2(SLC38A2) transporters.

Exercise is known to create a transient, but potent increase in skeletal muscle expression of potentially anti-inflammatory myokine interleukin-6 (IL-6). This effect may be clinically important in managing chronic inflammatory states. It has previously been proposed that lactic acidosis following exercise promotes this IL-6 up-regulation, but the mechanism of this acidosis effect is unknown. Rat skeletal muscle cell line L6-G8C5 has been used previously to model metabolic effects of acidosis, sensing low pH through the resulting inhibition of amino acid transporter SNAT2(SLC38A2). Use of ionophore ionomycin to model the rise in intracellular Ca2+ concentration occurring in contracting muscle strongly up-regulates IL-6 mRNA in L6-G8C5 myotubes. This study used this model to test the hypothesis that low extracellular pH (7.1) enhances ionomycin-induced IL-6 mRNA up-regulation by inhibiting SNAT2. Incubation of L6-G8C5 myotubes for 6 h with 0.5 µM ionomycin at control pH (7.4) resulted in a 15-fold increase in IL-6 mRNA which was further enhanced (1.74-fold) at pH 7.1. In contrast low pH had no significant effect on IL-6 mRNA without ionomycin, nor on the IL-6 mRNA increase that was induced by cyclic stretch. Even though pH 7.1 halved the transport activity of SNAT2, alternative methods of SNAT2 inhibition (JNK inhibitor SP600125; SNAT2 antagonist MeAIB; or SNAT2 silencing with siRNA) did not mimic the enhancing effect of low pH on IL-6 mRNA. On the contrary, JNK inhibition blunted the effect of pH 7.1 with ionomycin, but had no effect at pH 7.4. It is concluded that low pH promotes Ca2+/ionomycin-induced up-regulation of IL-6 mRNA through a novel SNAT2-independent JNK-dependent pH-sensing pathway not previously described in this skeletal muscle model.