3snd
From Proteopedia
Crystal structure of SARS coronavirus main protease complexed with Ac-ESTLQ-H (cocrystallization)
Structural highlights
Function[R1A_CVHSA] The papain-like proteinase (PL-PRO) is responsible for the cleavages located at the N-terminus of 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. Antagonizes innate immune induction of type I interferon by blocking the phosphorylation, dimerization and subsequent nuclear translocation of host IRF-3.[1] [2] [3] The main proteinase 3CL-PRO is responsible for the majority of cleavages as it cleaves the C-terminus of replicase polyprotein at 11 sites. Recognizes substrates containing the core sequence [ILMVF]-Q-|-[SGACN]. Inhibited by the substrate-analog Cbz-Val-Asn-Ser-Thr-Leu-Gln-CMK (By similarity). Also contains an ADP-ribose-1-phosphate (ADRP)-binding function.[4] [5] [6] Nsp7-nsp8 hexadecamer may possibly confer processivity to the polymerase, maybe by binding to dsRNA or by producing primers utilized by the latter.[7] [8] [9] Nsp9 is a ssRNA-binding protein.[10] [11] [12] Publication Abstract from PubMedSARS coronavirus main protease (SARS-CoV M(pro)) is essential for the replication of the virus and regarded as a major antiviral drug target. The enzyme is a cysteine protease, with a catalytic dyad (Cys-145/His-41) in the active site. Aldehyde inhibitors can bind reversibly to the active-site sulfhydryl of SARS-CoV M(pro). Previous studies using peptidic substrates and inhibitors showed that the substrate specificity of SARS-CoV M(pro) requires glutamine in the P1 position and a large hydrophobic residue in the P2 position. We determined four crystal structures of SARS-CoV M(pro) in complex with pentapeptide aldehydes (Ac-ESTLQ-H, Ac-NSFSQ-H, Ac-DSFDQ-H, and Ac-NSTSQ-H). Kinetic data showed that all of these aldehydes exhibit inhibitory activity towards SARS-CoV M(pro), with K(i) values in the muM range. Surprisingly, the X-ray structures revealed that the hydrophobic S2 pocket of the enzyme can accommodate serine and even aspartic-acid side-chains in the P2 positions of the inhibitors. Consequently, we reassessed the substrate specificity of the enzyme by testing the cleavage of 20 different tetradecapeptide substrates with varying amino-acid residues in the P2 position. The cleavage efficiency for the substrate with serine in the P2 position was 160-times lower than that for the original substrate (P2=Leu); furthermore, the substrate with aspartic acid in the P2 position was not cleaved at all. We also determined a crystal structure of SARS-CoV M(pro) in complex with aldehyde Cm-FF-H, which has its P1-phenylalanine residue bound to the relatively hydrophilic S1 pocket of the enzyme and yet exhibits a high inhibitory activity against SARS-CoV M(pro), with K(i)=2.24+/-0.58muM. These results show that the stringent substrate specificity of the SARS-CoV M(pro) with respect to the P1 and P2 positions can be overruled by the highly electrophilic character of the aldehyde warhead, thereby constituting a deviation from the dogma that peptidic inhibitors need to correspond to the observed cleavage specificity of the target protease. Peptide aldehyde inhibitors challenge the substrate specificity of the SARS-coronavirus main protease.,Zhu L, George S, Schmidt MF, Al-Gharabli SI, Rademann J, Hilgenfeld R Antiviral Res. 2011 Aug 11. PMID:21854807[13] From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine. See AlsoReferences
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