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10.1128/AAC.04698-14

http://scihub22266oqcxt.onion/10.1128/AAC.04698-14
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25421478!4356770!25421478
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suck abstract from ncbi


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pmid25421478      Antimicrob+Agents+Chemother 2015 ; 59 (4): 1827-36
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  • Biochemical characterization of recombinant Enterovirus 71 3C protease with fluorogenic model peptide substrates and development of a biochemical assay #MMPMID25421478
  • Shang L; Zhang S; Yang X; Sun J; Li L; Cui Z; He Q; Guo Y; Sun Y; Yin Z
  • Antimicrob Agents Chemother 2015[Apr]; 59 (4): 1827-36 PMID25421478show ga
  • Enterovirus 71 (EV71), a primary pathogen of hand, foot, and mouth disease (HFMD), affects primarily infants and children. Currently, there are no effective drugs against HFMD. EV71 3C protease performs multiple tasks in the viral replication, which makes it an ideal antiviral target. We synthesized a small set of fluorogenic model peptides derived from cleavage sites of EV71 polyprotein and examined their efficiencies of cleavage by EV71 3C protease. The novel peptide P08 [(2-(N-methylamino)benzoyl) (NMA)-IEALFQGPPK(DNP)FR] was determined to be the most efficiently cleaved by EV71 3C protease, with a kinetic constant kcat/Km of 11.8 +/- 0.82 mM(-1) min(-1). Compared with literature reports, P08 gave significant improvement in the signal/background ratio, which makes it an attractive substrate for assay development. A Molecular dynamics simulation study elaborated the interactions between substrate P08 and EV71 3C protease. Arg39, which is located at the bottom of the S2 pocket of EV71 3C protease, may participate in the proteolysis process of substrates. With an aim to evaluate EV71 3C protease inhibitors, a reliable and robust biochemical assay with a Z' factor of 0.87 +/- 0.05 was developed. A novel compound (compound 3) (50% inhibitory concentration [IC50] = 1.89 +/- 0.25 muM) was discovered using this assay, which effectively suppressed the proliferation of EV 71 (strain Fuyang) in rhabdomyosarcoma (RD) cells with a highly selective index (50% effective concentration [EC50] = 4.54 +/- 0.51 muM; 50% cytotoxic concentration [CC50] > 100 muM). This fast and efficient assay for lead discovery and optimization provides an ideal platform for anti-EV71 drug development targeting 3C protease.
  • |3C Viral Proteases[MESH]
  • |Amino Acid Sequence[MESH]
  • |Antiviral Agents/*chemical synthesis/*pharmacology[MESH]
  • |Biological Assay[MESH]
  • |Cell Line[MESH]
  • |Cell Survival/drug effects[MESH]
  • |Cysteine Endopeptidases/biosynthesis[MESH]
  • |Enterovirus/*enzymology[MESH]
  • |Fluorescent Dyes/*chemical synthesis[MESH]
  • |Foot-and-Mouth Disease Virus/*drug effects[MESH]
  • |Humans[MESH]
  • |Kinetics[MESH]
  • |Molecular Sequence Data[MESH]
  • |Peptides/*chemical synthesis/*pharmacology[MESH]
  • |Protease Inhibitors/*chemical synthesis/*pharmacology[MESH]
  • |Substrate Specificity[MESH]


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