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Discovery of a new Pro-Pro endopeptidase, PPEP-2, provides mechanistic insights into the differences in substrate specificity within the PPEP family.
Klychnikov, Oleg I; Shamorkina, Tatiana M; Weeks, Stephen D; van Leeuwen, Hans C; Corver, Jeroen; Drijfhout, Jan W; van Veelen, Peter A; Sluchanko, Nikolai N; Strelkov, Sergei V; Hensbergen, Paul J.
Afiliación
  • Klychnikov OI; From the Laboratory for Biocrystallography, Department of Pharmaceutical and Pharmacological Sciences, KU Leuven, 3000 Leuven, Belgium.
  • Shamorkina TM; the Center for Proteomics and Metabolomics and.
  • Weeks SD; From the Laboratory for Biocrystallography, Department of Pharmaceutical and Pharmacological Sciences, KU Leuven, 3000 Leuven, Belgium.
  • van Leeuwen HC; the Departments of Medical Microbiology and.
  • Corver J; the Departments of Medical Microbiology and.
  • Drijfhout JW; Immunohematology and Blood Transfusion, Leiden University Medical Center, 2300 Leiden, The Netherlands.
  • van Veelen PA; the Center for Proteomics and Metabolomics and.
  • Sluchanko NN; the A. N. Bach Institute of Biochemistry, Federal Research Center of Biotechnology of the Russian Academy of Sciences, 119071 Moscow, Russia, and.
  • Strelkov SV; the Department of Biophysics, Faculty of Biology, M. V. Lomonosov Moscow State University, 119991 Moscow, Russia.
  • Hensbergen PJ; From the Laboratory for Biocrystallography, Department of Pharmaceutical and Pharmacological Sciences, KU Leuven, 3000 Leuven, Belgium.
J Biol Chem ; 293(28): 11154-11165, 2018 07 13.
Article en En | MEDLINE | ID: mdl-29794027
Pro-Pro endopeptidases (PPEPs) belong to a recently discovered family of proteases capable of hydrolyzing a Pro-Pro bond. The first member from the bacterial pathogen Clostridium difficile (PPEP-1) cleaves two C. difficile cell-surface proteins involved in adhesion, one of which is encoded by the gene adjacent to the ppep-1 gene. However, related PPEPs may exist in other bacteria and may shed light on substrate specificity in this enzyme family. Here, we report on the homolog of PPEP-1 in Paenibacillus alvei, which we denoted PPEP-2. We found that PPEP-2 is a secreted metalloprotease, which likewise cleaved a cell-surface protein encoded by an adjacent gene. However, the cleavage motif of PPEP-2, PLP↓PVP, is distinct from that of PPEP-1 (VNP↓PVP). As a result, an optimal substrate peptide for PPEP-2 was not cleaved by PPEP-1 and vice versa. To gain insight into the specificity mechanism of PPEP-2, we determined its crystal structure at 1.75 Å resolution and further confirmed the structure in solution using small-angle X-ray scattering (SAXS). We show that a four-amino-acid loop, which is distinct in PPEP-1 and -2 (GGST in PPEP-1 and SERV in PPEP-2), plays a crucial role in substrate specificity. A PPEP-2 variant, in which the four loop residues had been swapped for those from PPEP-1, displayed a shift in substrate specificity toward PPEP-1 substrates. Our results provide detailed insights into the PPEP-2 structure and the structural determinants of substrate specificity in this new family of PPEP proteases.
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Texto completo: 1 Colección: 01-internacional Base de datos: MEDLINE Asunto principal: Endopeptidasas / Proteínas Bacterianas / Dipéptidos / Paenibacillus Tipo de estudio: Prognostic_studies Idioma: En Revista: J Biol Chem Año: 2018 Tipo del documento: Article País de afiliación: Bélgica Pais de publicación: Estados Unidos
Texto completo
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Texto completo: 1 Colección: 01-internacional Base de datos: MEDLINE Asunto principal: Endopeptidasas / Proteínas Bacterianas / Dipéptidos / Paenibacillus Tipo de estudio: Prognostic_studies Idioma: En Revista: J Biol Chem Año: 2018 Tipo del documento: Article País de afiliación: Bélgica Pais de publicación: Estados Unidos