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Biochemical and structural characterization of SplD protease from Staphylococcus aureus

Biochemical and structural characterization of SplD ...

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dc.contributor.author Zdżalik, Michał [USOS11798] pl
dc.contributor.author Kalińska, Magdalena [SAP14013626] pl
dc.contributor.author Wysocka, Magdalena pl
dc.contributor.author Steć-Niemczyk, Justyna [USOS57568] pl
dc.contributor.author Cichoń, Przemysław [USOS39539] pl
dc.contributor.author Stach, Natalia [USOS84126] pl
dc.contributor.author Gruba, Natalia pl
dc.contributor.author Stennicke, Henning R. pl
dc.contributor.author Jabaiah, Abeer pl
dc.contributor.author Markiewicz, Michał [SAP11019432] pl
dc.contributor.author Kędracka-Krok, Sylwia [SAP11016010] pl
dc.contributor.author Władyka, Benedykt [SAP12119116] pl
dc.contributor.author Daugherty, Patrick S. pl
dc.contributor.author Lesner, Adam pl
dc.contributor.author Rolka, Krzysztof pl
dc.contributor.author Dubin, Adam [SAP11006048] pl
dc.contributor.author Potempa, Jan [SAP11010833] pl
dc.contributor.author Dubin, Grzegorz [SAP11019243] pl
dc.date.accessioned 2015-04-23T08:37:47Z
dc.date.available 2015-04-23T08:37:47Z
dc.date.issued 2013 pl
dc.identifier.uri http://ruj.uj.edu.pl/xmlui/handle/item/5558
dc.language eng pl
dc.rights Udzielam licencji. Uznanie autorstwa 3.0 Polska *
dc.rights.uri http://creativecommons.org/licenses/by/3.0/pl/legalcode *
dc.title Biochemical and structural characterization of SplD protease from Staphylococcus aureus pl
dc.type JournalArticle pl
dc.abstract.en Staphylococcus aureus is a dangerous human pathogen. A number of the proteins secreted by this bacterium are implicated in its virulence, but many of the components of its secretome are poorly characterized. Strains of S. aureus can produce up to six homologous extracellular serine proteases grouped in a single spl operon. Although the SplA, SplB, and SplC proteases have been thoroughly characterized, the properties of the other three enzymes have not yet been investigated. Here, we describe the biochemical and structural characteristics of the SplD protease. The active enzyme was produced in an Escherichia coli recombinant system and purified to homogeneity. P1 substrate specificity was determined using a combinatorial library of synthetic peptide substrates showing exclusive preference for threonine, serine, leucine, isoleucine, alanine, and valine. To further determine the specificity of SplD, we used high-throughput synthetic peptide and cell surface protein display methods. The results not only confirmed SplD preference for a P1 residue, but also provided insight into the specificity of individual primed- and non-primed substrate-binding subsites. The analyses revealed a surprisingly narrow specificity of the protease, which recognized five consecutive residues (P4-P3-P2-P1-P1’) with a consensus motif of R-(Y/W)-(P/L)-(T/L/I/V)↓S. To understand the molecular basis of the strict substrate specificity, we crystallized the enzyme in two different conditions, and refined the structures at resolutions of 1.56 Å and 2.1 Å. Molecular modeling and mutagenesis studies allowed us to define a consensus model of substrate binding, and illustrated the molecular mechanism of protease specificity. pl
dc.subject.en serine protease pl
dc.subject.en infections pl
dc.subject.en proteinases pl
dc.subject.en chymotrypsin pl
dc.subject.en United States pl
dc.subject.en crystal structures pl
dc.subject.en nasal carriage pl
dc.subject.en extracellular proteases pl
dc.subject.en zymogen activation pl
dc.subject.en substrate specificity pl
dc.description.volume 8 pl
dc.description.number 10 pl
dc.identifier.doi 10.1371/journal.pone.0076812 pl
dc.identifier.eissn 1932-6203 pl
dc.title.journal PLoS ONE pl
dc.language.container eng pl
dc.affiliation Wydział Biochemii, Biofizyki i Biotechnologii : Zakład Mikrobiologii pl
dc.affiliation Wydział Biochemii, Biofizyki i Biotechnologii : Zakład Biochemii Analitycznej pl
dc.affiliation Wydział Biochemii, Biofizyki i Biotechnologii : Zakład Biochemii Fizycznej pl
dc.affiliation Wydział Biochemii, Biofizyki i Biotechnologii : Zakład Biofizyki Obliczeniowej i Bioinformatyki pl
dc.affiliation Pion Prorektora ds. badań naukowych i funduszy strukturalnych : Małopolskie Centrum Biotechnologii pl
dc.subtype Article pl
dc.identifier.articleid e76812 pl
dc.rights.original CC-BY; otwarte czasopismo; ostateczna wersja wydawcy; w momencie opublikowania; 0; pl
dc.identifier.project ROD UJ / P pl
dc.pbn.affiliation USOS84126:UJ.WBt; USOS57568:UJ.WBt; USOS39539:UJ.WBt; USOS11798:UJ.WBt; pl
.pointsMNiSW [2013 A]: 40


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Udzielam licencji. Uznanie autorstwa 3.0 Polska Except where otherwise noted, this item's license is described as Udzielam licencji. Uznanie autorstwa 3.0 Polska