Identification of Small Molecule Inhibitors against <i>Staphylococcus aureus</i> Dihydroorotase via HTS
Drug-resistant <i>Staphylococcus aureus</i> is an imminent threat to public health, increasing the importance of drug discovery utilizing unexplored bacterial pathways and enzyme targets. <i>De novo</i> pyrimidine biosynthesis is a specialized, highly conserved pathway implic...
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2021-09-01
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author | Amy J. Rice Russell P. Pesavento Jinhong Ren Isoo Youn Youngjin Kwon Kassapa Ellepola Chun-Tao Che Michael E. Johnson Hyun Lee |
author_facet | Amy J. Rice Russell P. Pesavento Jinhong Ren Isoo Youn Youngjin Kwon Kassapa Ellepola Chun-Tao Che Michael E. Johnson Hyun Lee |
author_sort | Amy J. Rice |
collection | DOAJ |
description | Drug-resistant <i>Staphylococcus aureus</i> is an imminent threat to public health, increasing the importance of drug discovery utilizing unexplored bacterial pathways and enzyme targets. <i>De novo</i> pyrimidine biosynthesis is a specialized, highly conserved pathway implicated in both the survival and virulence of several clinically relevant pathogens. Class I dihydroorotase (DHOase) is a separate and distinct enzyme present in gram positive bacteria (i.e., <i>S. aureus</i>, <i>B. anthracis</i>) that converts carbamoyl-aspartate (Ca-asp) to dihydroorotate (DHO)—an integral step in the <i>de novo</i> pyrimidine biosynthesis pathway. This study sets forth a high-throughput screening (HTS) of 3000 fragment compounds by a colorimetry-based enzymatic assay as a primary screen, identifying small molecule inhibitors of <i>S. aureus</i> DHOase (<i>Sa</i>DHOase), followed by hit validation with a direct binding analysis using surface plasmon resonance (SPR). Competition SPR studies of six hit compounds and eight additional analogs with the substrate Ca-asp determined the best compound to be a competitive inhibitor with a <i>K</i><sub>D</sub> value of 11 µM, which is 10-fold tighter than Ca-asp. Preliminary structure–activity relationship (SAR) provides the foundation for further structure-based antimicrobial inhibitor design against <i>S. aureus</i>. |
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spelling | doaj.art-b9635ad716f4444cb9a43c6709e654ac2023-11-22T13:30:41ZengMDPI AGInternational Journal of Molecular Sciences1661-65961422-00672021-09-012218998410.3390/ijms22189984Identification of Small Molecule Inhibitors against <i>Staphylococcus aureus</i> Dihydroorotase via HTSAmy J. Rice0Russell P. Pesavento1Jinhong Ren2Isoo Youn3Youngjin Kwon4Kassapa Ellepola5Chun-Tao Che6Michael E. Johnson7Hyun Lee8Center for Biomolecular Sciences, College of Pharmacy, University of Illinois at Chicago, Chicago, IL 60607, USACenter for Biomolecular Sciences, College of Pharmacy, University of Illinois at Chicago, Chicago, IL 60607, USACenter for Biomolecular Sciences, College of Pharmacy, University of Illinois at Chicago, Chicago, IL 60607, USADepartment of Pharmaceutical Sciences, College of Pharmacy, University of Illinois at Chicago, Chicago, IL 60612, USACenter for Biomolecular Sciences, College of Pharmacy, University of Illinois at Chicago, Chicago, IL 60607, USADepartment of Oral Biology, College of Dentistry, University of Illinois at Chicago, Chicago, IL 60607, USADepartment of Pharmaceutical Sciences, College of Pharmacy, University of Illinois at Chicago, Chicago, IL 60612, USACenter for Biomolecular Sciences, College of Pharmacy, University of Illinois at Chicago, Chicago, IL 60607, USACenter for Biomolecular Sciences, College of Pharmacy, University of Illinois at Chicago, Chicago, IL 60607, USADrug-resistant <i>Staphylococcus aureus</i> is an imminent threat to public health, increasing the importance of drug discovery utilizing unexplored bacterial pathways and enzyme targets. <i>De novo</i> pyrimidine biosynthesis is a specialized, highly conserved pathway implicated in both the survival and virulence of several clinically relevant pathogens. Class I dihydroorotase (DHOase) is a separate and distinct enzyme present in gram positive bacteria (i.e., <i>S. aureus</i>, <i>B. anthracis</i>) that converts carbamoyl-aspartate (Ca-asp) to dihydroorotate (DHO)—an integral step in the <i>de novo</i> pyrimidine biosynthesis pathway. This study sets forth a high-throughput screening (HTS) of 3000 fragment compounds by a colorimetry-based enzymatic assay as a primary screen, identifying small molecule inhibitors of <i>S. aureus</i> DHOase (<i>Sa</i>DHOase), followed by hit validation with a direct binding analysis using surface plasmon resonance (SPR). Competition SPR studies of six hit compounds and eight additional analogs with the substrate Ca-asp determined the best compound to be a competitive inhibitor with a <i>K</i><sub>D</sub> value of 11 µM, which is 10-fold tighter than Ca-asp. Preliminary structure–activity relationship (SAR) provides the foundation for further structure-based antimicrobial inhibitor design against <i>S. aureus</i>.https://www.mdpi.com/1422-0067/22/18/9984<i>S. aureus</i>antimicrobialdihydroorotaseClass I DHOaseinhibitors |
spellingShingle | Amy J. Rice Russell P. Pesavento Jinhong Ren Isoo Youn Youngjin Kwon Kassapa Ellepola Chun-Tao Che Michael E. Johnson Hyun Lee Identification of Small Molecule Inhibitors against <i>Staphylococcus aureus</i> Dihydroorotase via HTS International Journal of Molecular Sciences <i>S. aureus</i> antimicrobial dihydroorotase Class I DHOase inhibitors |
title | Identification of Small Molecule Inhibitors against <i>Staphylococcus aureus</i> Dihydroorotase via HTS |
title_full | Identification of Small Molecule Inhibitors against <i>Staphylococcus aureus</i> Dihydroorotase via HTS |
title_fullStr | Identification of Small Molecule Inhibitors against <i>Staphylococcus aureus</i> Dihydroorotase via HTS |
title_full_unstemmed | Identification of Small Molecule Inhibitors against <i>Staphylococcus aureus</i> Dihydroorotase via HTS |
title_short | Identification of Small Molecule Inhibitors against <i>Staphylococcus aureus</i> Dihydroorotase via HTS |
title_sort | identification of small molecule inhibitors against i staphylococcus aureus i dihydroorotase via hts |
topic | <i>S. aureus</i> antimicrobial dihydroorotase Class I DHOase inhibitors |
url | https://www.mdpi.com/1422-0067/22/18/9984 |
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