User talk:Daniel Hausaman

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==Use of Remdesivir as a treatment for COVID-19==
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|DATE=May 30, 2012
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|BAMBEDDOI=10.1002/bmb.20630
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==Bacterial RNA Polymerase: New Insights on a Fundamental Molecular Machine==
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===Introduction to RNAP===
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== Introduction to the SARS-CoV-2 genome and proteins ==
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The virus contains an unsegmented, single strand positive sense RNA genome. The genome is 29,981 base pairs long (Sanche et al 2020). The virus is surrounded by a lipid envelope containing the nucleocapsid. The RNA genome of SARS-CoV-2 is made up of of 14 open reading frames (ORF), which can encode a range of structural and non-structural proteins, these proteins directly and indirectly affect the virulence and replication of the virus (Astuti and Ysrafil, 2020). The genes that code for the non-structural proteins are translated firstly into Open reading frame 1a and Open reading frame 1b this forms two proteins known as pp1a and pp1ab. Due to a ribosomal frame shift the proteins are supplemented by protease enzymes known as papain-like proteases (PLpro) and a serine type Mpro protease. Cleavage takes place between pp1a and pp1ab to form non-structural proteins (nsps) 1–11 and 1–16 (Astuti and Ysrafil, 2020). Majority of the nsps form a Replicase-transcriptase complex (RTC). In addition to this RTC transcribes a genomic template of viral entry to negative-sense genes of both the viral genome and subgenomic RNA as intermediate products, this is followed by transcription to positive-sense mRNAs that can be mediated by RNA dependent RNA polymerase. The next step is translation, subgenomic proteins are translated into structural and accessories proteins such as M, S, and E proteins these are insulated in the endoplasmic reticulum and then moved to the endoplasmic reticulum-Golgi intermediate compartment (ERGIC) (Chen, et al 2020).
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'''[[RNA polymerase]]''' (RNAP) is a molecular machine that copies DNA into RNA and is found in every living organism. The bacterial RNAP complex consists of six subunits (ββ’α2ωσ) and three channels. RNAP initially binds to DNA at the promoter, forming the closed complex<ref name='genetics'>Snyder, L. & Champness, W. (2007). Molecular genetics of bacteria (3rd ed.). Washington, D.C.: ASM Press.</ref>. The DNA surrounding the promoter sequence unwinds and forms the open complex (http://www.pingrysmartteam.com/RPo/RPo.htm - please note that different nomenclature is used)<ref>2006 Pingry SMART Team: RNA Polymerase Holoenzyme Open Promoter Complex (Rpo) Jmol Tutorial</ref>. RNAP releases from the promoter and transitions into the elongation complex (EC). The EC moves along the template strand, adding ribonucleotides to the 3’ hydroxyl of the growing RNA transcript.
 
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This tutorial uses the β’ subunit of the RNAP elongation complex of ''Thermus thermophilus''. The β’ subunit contains structures crucial for transcription, including the sites for ribonucleotide addition and catalysis. Double-stranded DNA enters RNAP through the active site channel, while ribonucleotides (NTPs) enter through the secondary channel. As the downstream DNA (dwDNA) enters, it separates into the template and non-template strands. The template strand forms an approximately 90 degree kink in the active site channel. At the kink, one DNA base pair becomes available for NTP pairing and translocates to the +1 site. An NTP enters the active site and induces conformational change of the trigger loop into the trigger helix. The trigger helix forms a three-helical bundle with the bridge helix. This bundle changes dimensions of the active site and facilitates positioning of the NTP for addition to the growing RNA strand. Upon addition of the nucleotide, the dwDNA and RNA/DNA hybrid translocate through RNAP with stabilization from the rudder. The growing RNA strand is separated by the lid and exits RNAP through the exit channel. The DNA template strand rejoins the non-template strand as it exits the active site channel.
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== Introduction to Remdesivir as an antiviral drug ==
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Remdesivir is a drug that was initially developed for use against Ebola and has shown to be effective against coronaviruses such as SARS-CoV, MERS-CoV, SARS-CoV-2, and other coronaviruses.
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SARS-CoV-2 is the virus that is responsible for the disease COVID-19 which is a respiratory disease that can inflict severe damage to multiple major organs including the heart, brain, and lungs. SARS-Cov2 relies upon RNA-dependent RNA polymerase (RdRp) to replicate the viral genetic material. Drugs such as nucleoside analogues can be used to inhibit this RNA dependant RNA polymerase enzyme and stop viral replication.
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Remdesivir initially labeled GS-5734 is an adenosine triphosphate analogue, used as a broad-spectrum antiviral drug meaning it can be used to inhibit a wide range of viruses that rely upon RNA-dependent RNA polymerase for genomic replication.
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<qt>file=CrestUWMilwaukee2011-Video1.mp4|width=710|height=440|autoplay=false|controller=true|loop=false</qt><br>
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<qt>file=CrestUWMilwaukee2011-Video2.mp4|width=710|height=440|autoplay=false|controller=true|loop=false</qt><br>
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'''Q1. How does Remdesivir stop viral replication in COVID-19?'''
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----
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'''Q2. How can Remdesivir be used on other RNA viruses?'''
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===Learning Objectives===
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<Structure load='7BV2' size='350' frame='true' align='right' caption='Non-structural protein complex bound to the active form of Remdesivir' scene='Insert optional scene name here' />
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*Examine the bending of DNA in the active site channel
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==Structure==
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*Determine how ribonucleotides enter the active site
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Remdesivir triphosphate has the molecular formula C12H16N5O13P3 the compound is made up of a C-nucleoside, an aromatic amine, a nitrile, a pyrrolotriazine and an organic triphosphate.
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*Address how RNA polymerase discriminates between ribonucleotides and deoxyribonucleotides
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RNA dependant RNA Polymerase complex is an enzyme used in the replication and transcription of viral RNA. the structure of the protein is made up of an nsp12 core catalytic unit, an nsp7-nsp8 heterodimer, and an additional nsp8 subunit
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*Describe how ribonucleotide triphosphates (NTPs) are oriented correctly in the active site for catalysis
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On the right-hand side shows the three-dimensional interactive structure of the nsp12-nsp7-nsp8 complex also known as the RdRp bound to the template-primer RNA and the triphosphate form of remdesivir which is highlighted in blue to view remdesivir bound to the protein complex <scene name='86/867374/2/1'>Click here</scene>
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*Evaluate the conformational changes of the trigger loop
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*Describe how the trigger helix is involved in catalysis
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==Tutorial: β’ Subunit of ''Thermus thermophilus'' RNAP==
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== Mechanism ==
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<StructureSection load='2o5j' size='400' side='right' caption='' scene='β’ Subunit of Bacterial RNAP' scene='Beta-Prime_Subunit_of_Bacterial_RNA_Polymerase/2o5j_overview_1/1'>
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<font size='3'>'''DNA Translocation and the RNA/DNA Hybrid'''</font>
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<scene name='Beta-Prime_Subunit_of_Bacterial_RNA_Polymerase/2o5j_dna_1/1'>DNA</scene> in the active site channel provides the genetic information for RNA transcription. The active site channel is 27 Å wide and accommodates both <scene name='Beta-Prime_Subunit_of_Bacterial_RNA_Polymerase/2o5j_dwdna_1/1'>downstream DNA</scene> (dwDNA) and an <scene name='Beta-Prime_Subunit_of_Bacterial_RNA_Polymerase/2o5j_hybrid_1/1'>RNA/DNA hybrid</scene><ref>PMID: 10499798</ref>. The <scene name='Beta-Prime_Subunit_of_Bacterial_RNA_Polymerase/2o5j_template_1/3'>DNA template strand</scene> (blue) provides the complementary sequence for the RNA transcript and threads through the active site channel adjacent to the active site. The <scene name='Beta-Prime_Subunit_of_Bacterial_RNA_Polymerase/2o5j_nontemplate_1/2'>non-template strand</scene> (dark blue), or coding strand, is held away from the active site by the clamp helices and rudder.
 
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The template strand is kinked at the junction between the dwDNA and RNA/DNA hybrid<ref name='elongation'>PMID: 17581590</ref>. The lone unpaired acceptor template base at the <scene name='Beta-Prime_Subunit_of_Bacterial_RNA_Polymerase/2o5j_kink_1/2'>+1 site</scene> is located at the kink<ref name='elongation' />. The base pair at the <scene name='Beta-Prime_Subunit_of_Bacterial_RNA_Polymerase/2o5j_plus2site_1/2'>+2 site</scene> is distorted<ref name='elongation' />. Upstream of the kink is the <scene name='Beta-Prime_Subunit_of_Bacterial_RNA_Polymerase/2o5j_hybrid_1/1'>RNA/DNA hybrid</scene>. This hybrid structure is comprised of the template strand and the complementary RNA transcript connected by hydrogen bonds. The most recently formed hybrid bond is located at the <scene name='Beta-Prime_Subunit_of_Bacterial_RNA_Polymerase/2o5j_minus1site_1/2'>-1 site</scene><ref name='elongation' />.
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Once administered remdesivir requires metabolic activation, this takes place after the drug diffuses into a cell. Phosphoamidase (HINT1) and esterases CES1 and CTSA transform remdesivir into GS-441524 mono-phosphate. This is then phosphorylated again to produce the active triphosphate analog.
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The <scene name='Beta-Prime_Subunit_of_Bacterial_RNA_Polymerase/2o5j_rudder_1/2'>rudder</scene> (coral) stabilizes the dwDNA and upstream RNA/DNA hybrid with numerous sidechain interactions. Two <scene name='Beta-Prime_Subunit_of_Bacterial_RNA_Polymerase/2o5j_rudderarg_1/1'>arginine sidechains</scene> are shown contacting the dwDNA and RNA/DNA hybrid<ref name='elongation' />. The rudder meets the <scene name='Beta-Prime_Subunit_of_Bacterial_RNA_Polymerase/2o5j_clamp_1/2'>clamp helices</scene> (dark grey) which interact with the σ subunit of RNAP.
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Remdesivir is a polymerase inhibitor, there are 2 main categories for polymerase inhibitors these are known as nucleoside analogues and allosteric inhibitors. Nucleoside analogues resemble viral building blocks such as Adenosine triphosphate. One of the ways this inhibits viral replication is because of competitive inhibition between the nucleoside analogue and the viral RNA. To view the RNA highlighted in red <scene name='86/867374/2/1'>Click here</scene>
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Viral replication can also be reduced by the incorporation of incorrect nucleotides into the viral genome this can result in delayed chain termination. This is initiated by the formation of a phosphodiester bond, between the 3′ hydroxyl group of Remdesivir triphosphate and the next nucleotide resulting in the termination of the viral RNA synthesis at the position of 3 nucleotides.
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The upstream internal chamber can accommodate a 9-bp RNA/DNA hybrid<ref name='elongation' />. The RNA/DNA hybrid encounters the <scene name='Beta-Prime_Subunit_of_Bacterial_RNA_Polymerase/2o5j_lid_1/2'>lid</scene> (light green) that sterically blocks continued elongation<ref name='elongation' />. The lid facilitates <scene name='Beta-Prime_Subunit_of_Bacterial_RNA_Polymerase/2o5j_lidcleavage_1/1'>cleavage of the hydrogen bond</scene>, releasing the growing RNA transcript into the exit channel. As the bond is cleaved, the template strand moves one position upstream through the active site channel. This process is called translocation. This allows the lone <scene name='Beta-Prime_Subunit_of_Bacterial_RNA_Polymerase/2o5j_acceptor_1/4'>unpaired template nucleotide</scene> to move into the +1 site adjacent to the active site where nucleotide addition occurs<ref name='elongation' />.
 
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<font size='3'>'''Nucleotide Addition'''</font>
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----
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'''Q3. What type of polymerase inhibitor is Remdesivir categorised as?'''
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----
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'''Q4. Why does Remdesivir require metabolic activation?'''
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----
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'''Q5. How many phosphate groups does the activated form of Remdesivir contain?'''
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----
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'''Q6. What bond is formed that initiates the chain termination?'''
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The <scene name='Beta-Prime_Subunit_of_Bacterial_RNA_Polymerase/2o5j_secondary_1/1'>secondary channel</scene>, which is bordered by the <scene name='Beta-Prime_Subunit_of_Bacterial_RNA_Polymerase/2o5j_rim_1/2'>rim helices</scene> (very dark blue), forms the entrance for both ribonucleotides (NTPs) and deoxyribonucleotides (dNTPs) into RNAP. The secondary channel's dimensions are 15 x 20 Å, preventing the entrance of dsDNA<ref name='loading'>PMID: 17581591</ref>. Nucleotides are coupled to a magnesium ion (MgII) as they enter. <scene name='Beta-Prime_Subunit_of_Bacterial_RNA_Polymerase/2o5j_asn737_1/2'>Asn737</scene> (indigo) evaluates ribose hydroxyl groups in order to discriminate between NTPs and dNTPs<ref name='loading' />. The hydrogen bonds on the incoming NTP must complement those of the acceptor template base. As the NTPs enter, it is proposed that <scene name='Beta-Prime_Subunit_of_Bacterial_RNA_Polymerase/2o5j_metthr_1/3'>Met1238 and Thr1088</scene> (cyan and magenta) aid in selection and orientation of the cognate NTP<ref name='loading' />. Once the cognate NTP is selected, its alpha and gamma phosphates form temporary phosphate contacts with RNAP near the active site<ref name='loading' />. The nucleotide adopts a relaxed conformation and resists loading into the catalytic position<ref name='loading' />. This conformation is influenced by the "basic rim gate" consisting of four residues that surround the NTP phosphates<ref name='loading' />. Two basic rim gate residues, <scene name='Beta-Prime_Subunit_of_Bacterial_RNA_Polymerase/2o5j_basicgate_1/2'>Arg783 and Arg1029</scene>, are shown (light purple).
 
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The <scene name='Beta-Prime_Subunit_of_Bacterial_RNA_Polymerase/2o5j_bridge_1/3'>bridge helix</scene> (magenta) separates the active and secondary channels while interacting with the <scene name='Beta-Prime_Subunit_of_Bacterial_RNA_Polymerase/2o5j_trigger_1/2'>trigger helix</scene> (cyan). In the pre-insertion state, the trigger loop has an unstructured conformation<ref name='loading' />. Loading of the NTP near the active site induces a conformational change in the trigger loop, and it becomes the two-helical trigger helix<ref name='loading' />. The <scene name='Beta-Prime_Subunit_of_Bacterial_RNA_Polymerase/2o5j_trigger2_1/1'>trigger helix</scene> "swings" into the secondary channel and changes the dimensions of the channel to 11 x 11 Å<ref name='loading' />. This reduction in size prevents diffusion of NTP away from the active site while simultaneously preventing interference from other nucleotides<ref name='loading' />. The presence of the trigger helix causes the NTP phosphate contacts to change<ref name='loading' />. All three phosphates contact RNAP and adopt a more rigid conformation<ref name='loading' />. The <scene name='Beta-Prime_Subunit_of_Bacterial_RNA_Polymerase/2o5j_ntp_1/2'>NTP</scene> is now ready for catalysis.
 
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<font size='3'>'''Catalysis'''</font>
 
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The <scene name='Beta-Prime_Subunit_of_Bacterial_RNA_Polymerase/2o5j_activesite_1/4'>active site</scene> consists of three highly conserved aspartate sidechains (Asp739, Asp741, Asp743) chelated to a magnesium ion (MgI) required for catalysis<ref name='loading' />. <scene name='Beta-Prime_Subunit_of_Bacterial_RNA_Polymerase/2o5j_mgions_1/2'>MgI and MgII</scene> (lime green) chelated to the NTP coordinate positioning of the NTP for catalysis<ref name='lehninger'>Nelson, D. L. & Cox, M. M. (2008). Lehninger principles of biochemistry (5th ed.). New York: W. H. Freeman and Company.
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== References ==
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</ref>. The <scene name='Beta-Prime_Subunit_of_Bacterial_RNA_Polymerase/2o5j_threeprime_1/2'>3' hydroxyl</scene> of the RNA transcript has nucleophilic activity and attacks the alpha-phosphate of the NTP<ref name='lehninger' />. A phosphodiester bond forms between the RNA transcript and the alpha-phosphate, and the beta- and gamma-phosphates leave as a pyrophosphate group<ref name='lehninger' />. After catalysis the RNA/DNA hybrid moves in the <scene name='Beta-Prime_Subunit_of_Bacterial_RNA_Polymerase/2o5j_minus1site_1/2'>-1 site</scene>, and the ribonucleotide in this bond provides the 3’ hydroxyl for the next incoming NTP<ref name='elongation' />.
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</StructureSection>
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===Nucleotide Addition Cycle===
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Hanna R, Dalvi S. (2021). Understanding COVID-19 Pandemic: Molecular Mechanisms and Potential Therapeutic Strategies. An Evidence-Based Review. Journal of Inflammation Research » Volume 14. https://doi.org/10.2147/JIR.S282213.
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Masters P. (2006) The molecular biology of coronaviruses. Adv Virus Res; 66:193-292. doi: 10.1016/S0065-3527(06)66005-3.
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<qt>file=CrestUWMilwaukee2011-Video3.mp4|width=710|height=440|autoplay=false|controller=true|loop=false</qt><br>
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Chen Y et al. (2020). Emerging coronaviruses: Genome structure, replication, and pathogenesis. J Med Virol; 92(4):418-423. doi: 10.1002/jmv.25681
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Greenwood M. (2021). Screening for SARS-CoV-2 non-structural protein 14 inhibitors. Available at: https://www.news-medical.net/news/20210412/Screening-for-SARS-CoV-2-non-structural-protein-14-inhibitors.aspx (Accessed 8th May 2021)
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An animation showing the conformational changes undergone by the trigger loop/helix when switching from the pre-insertion complex to the insertion complex can currently be found at http://www.molmovdb.org/cgi-bin/morph.cgi?ID=807081-19674. This animation was designed by Mark Hoelzer of the Center for BioMolecular Modeling at MSOE. The conformational change animation is an interpretation of static models, but does not represent the actual conformational change.
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Jinsung Y et al. (2020). Molecular interaction and inhibition of SARS-CoV-2 binding to the ACE2 receptor Nature Communications volume 11, Article number: 4541. https://doi.org/10.1038/s41467-020-18319-6
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Protein Database Bank in Europe 7bv2 › F86. Available at: https://www.ebi.ac.uk/pdbe/entry/pdb/7bv2/bound/F86 (Accessed: 12th June 2021)
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===Challenge Questions===
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Uzunova K et al. (2020). Insights into antiviral mechanisms of remdesivir, lopinavir/ritonavir and chloroquine/hydroxychloroquine affecting the new SARS-CoV-2. Biomed Pharmacother. 2020 Nov; 131: 110668. doi: 10.1016/j.biopha.2020.110668
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*Why does DNA experience a 90 degree bend in the active site channel?
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Yu-chen et al. (2020).Remdesivir for severe acute respiratory syndrome coronavirus2 causing COVID-19: An evaluation of the evidence. Travel Medicine and Infectious Disease. Volume 35, https://doi.org/10.1016/j.tmaid.2020.101647
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*What are the functions of the magnesium ion in the active site and the magnesium ion coupled to the incoming NTP? What are they coordinating?
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Olena A. (2020). Remdesivir Works Against Coronaviruses in the Lab. Available at: https://www.the-scientist.com/news-opinion/remdesivir-works-great-against-coronaviruses-in-the-lab-67298 (Accessed: 6th July 2021)
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*What experiments could prove the ribonucleotide discrimination function of β'Asn737?
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PubChem. Remdesivir. Available at: https://pubchem.ncbi.nlm.nih.gov/compound/Remdesivir (Accessed: 12 April 2021)
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*What experimental evidence could confirm that the trigger loop to trigger helix conformational change is involved in catalysis?
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Ghandi Z et al. (2020). Potential Interactions of Remdesivir with Pulmonary Drugs: a Covid-19 Perspective. SN Compr Clin Med: 1–2. doi: 10.1007/s42399-020-00462-2
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PubChem. Adenosine-5'-triphosphate Available at: https://pubchem.ncbi.nlm.nih.gov/compound/Adenosine-5_-triphosphate (Accessed 2nd March
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==2011 UW-Milwaukee CREST Team==
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===Team Members===
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Catherine L Dornfeld, Christopher Hanna and Jason Slaasted
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===Abstract===
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RNA polymerase (RNAP) is an information-processing molecular machine that copies DNA into RNA. It is a multi-subunit complex found in every living organism. Bacterial RNAP contains six subunits (ββ’α2ωσ). This model focuses on the β’ subunit of RNAP elongation complex (EC) of ''Thermus thermophilus'' that contains the active site sequence and several structures involved in the catalytic mechanism: the aspartate residues, the magnesium ions, the bridge helix, and the trigger helix. The active site channel accommodates double stranded DNA (dwDNA) and an RNA/DNA hybrid. The secondary channel, which is bordered by the rim helices, allows nucleotides (NTPs) to enter the active site. The exit channel guides the growing RNA transcript out of the complex. The DNA template strand becomes kinked as it moves through the active site channel and is separated from the non-template strand. This kink allows one dNTP at a time to become available for nucleotide addition once it translocates to the +1 site. The bridge helix (BH) and trigger loop (TL) work together as a “swinging gate” to enhance the catalytic action by facilitating NTP addition. In the crystal structure of the EC without NTP in the active site, the TL (β’ 1236-1265) is unstructured. In the EC crystal structure with a non-hydrolysable nucleotide (AMPcPP), the TL folds into two anti-parallel helices (trigger helix, TH) that interact with the adjacent BH to create a three-helical bundle forming a catalytically active complex. The other structures that are functionally important in the β’ subunit are the lid (β’ 525-539) that cleaves the RNA/DNA hybrid, directing the newly formed RNA out through the exit channel, and the rudder(β’ 582-602) that helps to stabilize the DNA helix and the RNA/DNA hybrid in the active site channel. The clamp helices interact with the σ subunit of RNAP.
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===Poster===
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[[Image:UWMFinal.jpg|Catherine L Dornfeld| 350px]]
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===Acknowledgments===
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*Steven Forst, Ph.D., University of Wisconsin-Milwaukee
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*Rick Gourse, Ph.D., University of Wisconsin-Madison
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*MSOE Center for BioMolecular Modeling: Mark Hoelzer, Margaret Franzen, Ph.D. and Tim Herman, Ph.D.
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*NSF CREST Program
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==References==
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<references />
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[[Category:Featured in BAMBED]]
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Current revision

Contents

Use of Remdesivir as a treatment for COVID-19

Introduction to the SARS-CoV-2 genome and proteins

The virus contains an unsegmented, single strand positive sense RNA genome. The genome is 29,981 base pairs long (Sanche et al 2020). The virus is surrounded by a lipid envelope containing the nucleocapsid. The RNA genome of SARS-CoV-2 is made up of of 14 open reading frames (ORF), which can encode a range of structural and non-structural proteins, these proteins directly and indirectly affect the virulence and replication of the virus (Astuti and Ysrafil, 2020). The genes that code for the non-structural proteins are translated firstly into Open reading frame 1a and Open reading frame 1b this forms two proteins known as pp1a and pp1ab. Due to a ribosomal frame shift the proteins are supplemented by protease enzymes known as papain-like proteases (PLpro) and a serine type Mpro protease. Cleavage takes place between pp1a and pp1ab to form non-structural proteins (nsps) 1–11 and 1–16 (Astuti and Ysrafil, 2020). Majority of the nsps form a Replicase-transcriptase complex (RTC). In addition to this RTC transcribes a genomic template of viral entry to negative-sense genes of both the viral genome and subgenomic RNA as intermediate products, this is followed by transcription to positive-sense mRNAs that can be mediated by RNA dependent RNA polymerase. The next step is translation, subgenomic proteins are translated into structural and accessories proteins such as M, S, and E proteins these are insulated in the endoplasmic reticulum and then moved to the endoplasmic reticulum-Golgi intermediate compartment (ERGIC) (Chen, et al 2020).


Introduction to Remdesivir as an antiviral drug

Remdesivir is a drug that was initially developed for use against Ebola and has shown to be effective against coronaviruses such as SARS-CoV, MERS-CoV, SARS-CoV-2, and other coronaviruses. SARS-CoV-2 is the virus that is responsible for the disease COVID-19 which is a respiratory disease that can inflict severe damage to multiple major organs including the heart, brain, and lungs. SARS-Cov2 relies upon RNA-dependent RNA polymerase (RdRp) to replicate the viral genetic material. Drugs such as nucleoside analogues can be used to inhibit this RNA dependant RNA polymerase enzyme and stop viral replication. Remdesivir initially labeled GS-5734 is an adenosine triphosphate analogue, used as a broad-spectrum antiviral drug meaning it can be used to inhibit a wide range of viruses that rely upon RNA-dependent RNA polymerase for genomic replication.

Q1. How does Remdesivir stop viral replication in COVID-19?

Q2. How can Remdesivir be used on other RNA viruses?

Non-structural protein complex bound to the active form of Remdesivir

Drag the structure with the mouse to rotate

Structure

Remdesivir triphosphate has the molecular formula C12H16N5O13P3 the compound is made up of a C-nucleoside, an aromatic amine, a nitrile, a pyrrolotriazine and an organic triphosphate. RNA dependant RNA Polymerase complex is an enzyme used in the replication and transcription of viral RNA. the structure of the protein is made up of an nsp12 core catalytic unit, an nsp7-nsp8 heterodimer, and an additional nsp8 subunit On the right-hand side shows the three-dimensional interactive structure of the nsp12-nsp7-nsp8 complex also known as the RdRp bound to the template-primer RNA and the triphosphate form of remdesivir which is highlighted in blue to view remdesivir bound to the protein complex

Mechanism

Once administered remdesivir requires metabolic activation, this takes place after the drug diffuses into a cell. Phosphoamidase (HINT1) and esterases CES1 and CTSA transform remdesivir into GS-441524 mono-phosphate. This is then phosphorylated again to produce the active triphosphate analog.

Remdesivir is a polymerase inhibitor, there are 2 main categories for polymerase inhibitors these are known as nucleoside analogues and allosteric inhibitors. Nucleoside analogues resemble viral building blocks such as Adenosine triphosphate. One of the ways this inhibits viral replication is because of competitive inhibition between the nucleoside analogue and the viral RNA. To view the RNA highlighted in red Viral replication can also be reduced by the incorporation of incorrect nucleotides into the viral genome this can result in delayed chain termination. This is initiated by the formation of a phosphodiester bond, between the 3′ hydroxyl group of Remdesivir triphosphate and the next nucleotide resulting in the termination of the viral RNA synthesis at the position of 3 nucleotides.


Q3. What type of polymerase inhibitor is Remdesivir categorised as?

Q4. Why does Remdesivir require metabolic activation?

Q5. How many phosphate groups does the activated form of Remdesivir contain?

Q6. What bond is formed that initiates the chain termination?



References

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