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caspase, substrates, article, substrate, pubmed, google, scholar, cas, doi, specificity, quenched, concentration, acc, internally, caspases, glu, enzyme, nature, pair, protease, iqf, mmp, fluorescence, trypsin, proteases, mca, cleavage, peptide, amino, cell, activity, residues, assay, enzymes, asp, library, libraries, acids, size, buffer, analysis, human, kinetic, central, full, gly, ranged, preferences, fluorophore, elastase,

Topics {✒️}

nature portfolio privacy policy 7-methoxy-coumarin-4-yl acetic acid-2 author information authors full-length substrate acc-gdevdgvk membrane-bound collagenase mt1-mmp advertising general formula acc-gglsxivk social media canada research chair chemical tools hydrophobic nature nature methods 12 nature methods 2 protease-based drug discovery 0/ reprints nature nature 410 izabela rutkowska-wlodarczyk atherosclerosis-based cardiovascular disease conserved xiap-interaction motif caspase substrate acc-gdevdgvk horizon 2020 research blood research designed research p1-asp/glu substrate libraries solid-phase library synthesis excellent tools full size image performed research synthesizing fluorophore-gdevdgvk 1 μm solutions develop substrate-based markers50 p1-p3 peptide libraries solid-phase peptide synthesis itraq-tails quantitative proteomics interleukin-1-beta converting-enzyme multiplex n-terminome analysis pair comprises 7-amino-4-carbamoylmethylcoumarin post-translationally modified residues adaptable donor/acceptor pair post-translationally modified lysine post-translationally modified peptides apoptotic cell-based experiments post-translationally modified arginine donor/acceptor pair acc/dnp acc/dnp donor/acceptor pair d-amino acid inhibitors full size table c-terminal glu residues45

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• Matrix metalloproteinases: what do they not do?

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         description:Internally quenched fluorescent (IQF) peptide substrates originating from FRET (Förster Resonance Energy Transfer) are powerful tool for examining the activity and specificity of proteases, and a variety of donor/acceptor pairs are extensively used to design individual substrates and combinatorial libraries. We developed a highly sensitive and adaptable donor/acceptor pair that can be used to investigate the substrate specificity of cysteine proteases, serine proteases and metalloproteinases. This novel pair comprises 7-amino-4-carbamoylmethylcoumarin (ACC) as the fluorophore and 2,4-dinitrophenyl-lysine (Lys(DNP)) as the quencher. Using caspase-3, caspase-7, caspase-8, neutrophil elastase, legumain, and two matrix metalloproteinases (MMP2 and MMP9), we demonstrated that substrates containing ACC/Lys(DNP) exhibit 7 to 10 times higher sensitivity than conventional 7-methoxy-coumarin-4-yl acetic acid (MCA)/Lys(DNP) substrates; thus, substantially lower amounts of substrate and enzyme can be used for each assay. We therefore propose that the ACC/Lys(DNP) pair can be considered a novel and sensitive scaffold for designing substrates for any group of endopeptidases. We further demonstrate that IQF substrates containing unnatural amino acids can be used to investigate protease activities/specificities for peptides containing post-translationally modified amino acids. Finally, we used IQF substrates to re-investigate the P1-Asp characteristic of caspases, thus demonstrating that some human caspases can also hydrolyze substrates after glutamic acid.
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      affiliation:
            name:Faculty of Chemistry, Wroclaw University of Technology, Wyb
            address:
               name:Department of Bioorganic Chemistry, Faculty of Chemistry, Wroclaw University of Technology, Wyb, Wyspianskiego 27, Poland
               type:PostalAddress
            type:Organization
      name:Paulina Kasperkiewicz
      affiliation:
            name:Faculty of Chemistry, Wroclaw University of Technology, Wyb
            address:
               name:Department of Bioorganic Chemistry, Faculty of Chemistry, Wroclaw University of Technology, Wyb, Wyspianskiego 27, Poland
               type:PostalAddress
            type:Organization
            name:NCI-designated Cancer Center, Sanford-Burnham Prebys Medical Discovery Institute, La Jolla
            address:
               name:NCI-designated Cancer Center, Sanford-Burnham Prebys Medical Discovery Institute, La Jolla, USA
               type:PostalAddress
            type:Organization
      name:Izabela Rutkowska-Wlodarczyk
      affiliation:
            name:Laboratory of Neurobiology, Nencki Institute of Experimental Biology, Polish Academy of Sciences
            address:
               name:Laboratory of Neurobiology, Nencki Institute of Experimental Biology, Polish Academy of Sciences, Poland
               type:PostalAddress
            type:Organization
      name:Scott J. Snipas
      affiliation:
            name:NCI-designated Cancer Center, Sanford-Burnham Prebys Medical Discovery Institute, La Jolla
            address:
               name:NCI-designated Cancer Center, Sanford-Burnham Prebys Medical Discovery Institute, La Jolla, USA
               type:PostalAddress
            type:Organization
      name:Yoshifumi Itoh
      affiliation:
            name:Kennedy Institute of Rheumatology, University of Oxford
            address:
               name:Kennedy Institute of Rheumatology, University of Oxford, United Kingdom
               type:PostalAddress
            type:Organization
      name:Dusan Turk
      affiliation:
            name:Jožef Stefan Institute
            address:
               name:Department of Biochemistry and Molecular and Structural Biology, Jožef Stefan Institute, Slovenia
               type:PostalAddress
            type:Organization
      name:Boris Turk
      affiliation:
            name:Jožef Stefan Institute
            address:
               name:Department of Biochemistry and Molecular and Structural Biology, Jožef Stefan Institute, Slovenia
               type:PostalAddress
            type:Organization
      name:Christopher M. Overall
      affiliation:
            name:Centre for Blood Research, University of British Columbia
            address:
               name:Department of Oral Biological and Medical Sciences, Centre for Blood Research, University of British Columbia, Vancouver, Canada
               type:PostalAddress
            type:Organization
      name:Leszek Kaczmarek
      affiliation:
            name:Laboratory of Neurobiology, Nencki Institute of Experimental Biology, Polish Academy of Sciences
            address:
               name:Laboratory of Neurobiology, Nencki Institute of Experimental Biology, Polish Academy of Sciences, Poland
               type:PostalAddress
            type:Organization
      name:Guy S. Salvesen
      affiliation:
            name:NCI-designated Cancer Center, Sanford-Burnham Prebys Medical Discovery Institute, La Jolla
            address:
               name:NCI-designated Cancer Center, Sanford-Burnham Prebys Medical Discovery Institute, La Jolla, USA
               type:PostalAddress
            type:Organization
      email:[email protected]
      name:Marcin Drag
      affiliation:
            name:Faculty of Chemistry, Wroclaw University of Technology, Wyb
            address:
               name:Department of Bioorganic Chemistry, Faculty of Chemistry, Wroclaw University of Technology, Wyb, Wyspianskiego 27, Poland
               type:PostalAddress
            type:Organization
      email:[email protected]
PostalAddress:
      name:Department of Bioorganic Chemistry, Faculty of Chemistry, Wroclaw University of Technology, Wyb, Wyspianskiego 27, Poland
      name:NCI-designated Cancer Center, Sanford-Burnham Prebys Medical Discovery Institute, La Jolla, USA
      name:Department of Bioorganic Chemistry, Faculty of Chemistry, Wroclaw University of Technology, Wyb, Wyspianskiego 27, Poland
      name:Department of Bioorganic Chemistry, Faculty of Chemistry, Wroclaw University of Technology, Wyb, Wyspianskiego 27, Poland
      name:Department of Bioorganic Chemistry, Faculty of Chemistry, Wroclaw University of Technology, Wyb, Wyspianskiego 27, Poland
      name:NCI-designated Cancer Center, Sanford-Burnham Prebys Medical Discovery Institute, La Jolla, USA
      name:Laboratory of Neurobiology, Nencki Institute of Experimental Biology, Polish Academy of Sciences, Poland
      name:NCI-designated Cancer Center, Sanford-Burnham Prebys Medical Discovery Institute, La Jolla, USA
      name:Kennedy Institute of Rheumatology, University of Oxford, United Kingdom
      name:Department of Biochemistry and Molecular and Structural Biology, Jožef Stefan Institute, Slovenia
      name:Department of Biochemistry and Molecular and Structural Biology, Jožef Stefan Institute, Slovenia
      name:Department of Oral Biological and Medical Sciences, Centre for Blood Research, University of British Columbia, Vancouver, Canada
      name:Laboratory of Neurobiology, Nencki Institute of Experimental Biology, Polish Academy of Sciences, Poland
      name:NCI-designated Cancer Center, Sanford-Burnham Prebys Medical Discovery Institute, La Jolla, USA
      name:Department of Bioorganic Chemistry, Faculty of Chemistry, Wroclaw University of Technology, Wyb, Wyspianskiego 27, Poland

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