4yoi | pdb_00004yoi
Structure of HKU4 3CLpro bound to non-covalent inhibitor 1A
Structural highlights
FunctionR1AB_BCHK4 The replicase polyprotein of coronaviruses is a multifunctional protein: it contains the activities necessary for the transcription of negative stranded RNA, leader RNA, subgenomic mRNAs and progeny virion RNA as well as proteinases responsible for the cleavage of the polyprotein into functional products.[UniProtKB:P0C6X7] Host translation inhibitor nsp1: Inhibits host translation by interacting with the 40S ribosomal subunit. The nsp1-40S ribosome complex further induces an endonucleolytic cleavage near the 5'UTR of host mRNAs, targeting them for degradation. Viral mRNAs are not susceptible to nsp1-mediated endonucleolytic RNA cleavage thanks to the presence of a 5'-end leader sequence and are therefore protected from degradation. By suppressing host gene expression, nsp1 facilitates efficient viral gene expression in infected cells and evasion from host immune response.[UniProtKB:P0C6X7] Non-structural protein 2: May play a role in the modulation of host cell survival signaling pathway by interacting with host PHB and PHB2. Indeed, these two proteins play a role in maintaining the functional integrity of the mitochondria and protecting cells from various stresses.[UniProtKB:P0C6X7] Papain-like proteinase: Responsible for the cleavages located at the N-terminus of the replicase polyprotein. In addition, PL-PRO possesses a deubiquitinating/deISGylating activity and processes both 'Lys-48'- and 'Lys-63'-linked polyubiquitin chains from cellular substrates. Participates together with nsp4 in the assembly of virally-induced cytoplasmic double-membrane vesicles necessary for viral replication. Antagonizes innate immune induction of type I interferon by blocking the phosphorylation, dimerization and subsequent nuclear translocation of host IRF3. Prevents also host NF-kappa-B signaling.[UniProtKB:P0C6X7] Non-structural protein 4: Participates in the assembly of virally-induced cytoplasmic double-membrane vesicles necessary for viral replication.[UniProtKB:P0C6X7] Proteinase 3CL-PRO: Cleaves the C-terminus of replicase polyprotein at 11 sites. Recognizes substrates containing the core sequence [ILMVF]-Q-|-[SGACN]. Also able to bind an ADP-ribose-1-phosphate (ADRP).[UniProtKB:P0C6X7][PROSITE-ProRule:PRU00772] Non-structural protein 6: Plays a role in the initial induction of autophagosomes from host reticulum endoplasmic. Later, limits the expansion of these phagosomes that are no longer able to deliver viral components to lysosomes.[UniProtKB:P0C6X7] Non-structural protein 7: Forms an hexadecamer with nsp8 (8 subunits of each) that may participate in viral replication by acting as a primase. Alternatively, may synthesize substantially longer products than oligonucleotide primers.[UniProtKB:P0C6X7] Non-structural protein 8: Forms an hexadecamer with nsp7 (8 subunits of each) that may participate in viral replication by acting as a primase. Alternatively, may synthesize substantially longer products than oligonucleotide primers.[UniProtKB:P0C6X7] Non-structural protein 9: May participate in viral replication by acting as a ssRNA-binding protein.[UniProtKB:P0C6X7] Non-structural protein 10: Plays a pivotal role in viral transcription by stimulating both nsp14 3'-5' exoribonuclease and nsp16 2'-O-methyltransferase activities. Therefore plays an essential role in viral mRNAs cap methylation.[UniProtKB:P0C6X7] RNA-directed RNA polymerase: Responsible for replication and transcription of the viral RNA genome.[UniProtKB:P0C6X7] Helicase: Multi-functional protein with a zinc-binding domain in N-terminus displaying RNA and DNA duplex-unwinding activities with 5' to 3' polarity. Activity of helicase is dependent on magnesium.[UniProtKB:P0C6X7] Guanine-N7 methyltransferase: Enzyme possessing two different activities: an exoribonuclease activity acting on both ssRNA and dsRNA in a 3' to 5' direction and a N7-guanine methyltransferase activity.[UniProtKB:P0C6X7] Uridylate-specific endoribonuclease: Mn(2+)-dependent, uridylate-specific enzyme, which leaves 2'-3'-cyclic phosphates 5' to the cleaved bond.[UniProtKB:P0C6X7] 2'-O-methyltransferase: Methyltransferase that mediates mRNA cap 2'-O-ribose methylation to the 5'-cap structure of viral mRNAs. N7-methyl guanosine cap is a prerequisite for binding of nsp16. Therefore plays an essential role in viral mRNAs cap methylation which is essential to evade immune system.[UniProtKB:P0C6X7] Publication Abstract from PubMedThe bat coronavirus HKU4 belongs to the same 2c lineage as that of the deadly Middle East Respiratory Syndrome coronavirus (MERS-CoV) and shows high sequence similarity, therefore potentiating a threat to the human population through a zoonotic shift or 'spill over' event. To date, there are no effective vaccines or antiviral treatments available that are capable of limiting the pathogenesis of any human coronaviral infection. An attractive target for the development of anti-coronaviral therapeutics is the 3C-like protease (3CLpro), which is essential for the progression of the coronaviral life cycle. Herein, we report the screening results of a small, 230-member peptidomimetic library against HKU4-CoV 3CLpro and the identification of 43 peptidomimetic compounds showing good to excellent inhibitory potency of HKU4-CoV 3CLpro with IC50 values ranging from low micromolar to sub-micromolar. We established structure-activity relationships (SARs) describing the important ligand-based features required for potent HKU4-CoV 3CLpro inhibition and identified a seemingly favored peptidic backbone for HKU4-CoV 3CLpro inhibition. To investigate this, a molecular sub-structural analysis of the most potent HKU4-CoV 3CLpro inhibitor was accomplished by the synthesis and testing of the lead peptidomimetic inhibitor's sub-structural components, confirming the activity of the favored backbone (22A) identified via SAR analysis. In order to elucidate the structural reasons for such potent HKU4-CoV 3CLpro inhibition by the peptidomimetics having the 22A backbone, we determined the X-ray structures of HKU4-CoV 3CLpro in complex with three peptidomimetic inhibitors. Sequence alignment of HKU4-CoV 3CLpro, and two other lineage C Betacoronaviruses 3CLpro's, HKU5-CoV and MERS-CoV 3CLpro, show that the active site residues of HKU4-CoV 3CLpro that participate in inhibitor binding are conserved in HKU5-CoV and MERS-CoV 3CLpro. Furthermore, we assayed our most potent HKU4-CoV 3CLpro inhibitor for inhibition of HKU5-CoV 3CLpro and found it to have sub-micromolar inhibitory activity (IC50=0.54+/-0.03muM). The X-ray structures and SAR analysis reveal critical insights into the structure and inhibition of HKU4-CoV 3CLpro, providing fundamental knowledge that may be exploited in the development of anti-coronaviral therapeutics for coronaviruses emerging from zoonotic reservoirs. Targeting zoonotic viruses: Structure-based inhibition of the 3C-like protease from bat coronavirus HKU4-The likely reservoir host to the human coronavirus that causes Middle East Respiratory Syndrome (MERS).,St John SE, Tomar S, Stauffer SR, Mesecar AD Bioorg Med Chem. 2015 Jun 19. pii: S0968-0896(15)00533-7. doi:, 10.1016/j.bmc.2015.06.039. PMID:26190463[1] From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine. References
| ||||||||||||||||||
