title: Capsule-Targeting Depolymerase, Derived from Klebsiella KP36 Phage, as a Tool for the Development of Anti-Virulent Strategy.
abstract: The rise of antibiotic-resistantKlebsiella pneumoniae, a leading nosocomial pathogen, prompts the need for alternative therapies. We have identified and characterized a novel depolymerase enzyme encoded byKlebsiellaphage KP36 (depoKP36), from theSiphoviridaefamily. To gain insights into the catalytic and structural features of depoKP36, we have recombinantly produced this protein of 93.4 kDa and showed that it is able to hydrolyze a crude exopolysaccharide of aK. pneumoniaehost. Using in vitro and in vivo assays, we found that depoKP36 was also effective against a native capsule of clinicalK. pneumoniaestrains, representing the K63 type, and significantly inhibitedKlebsiella-induced mortality ofGalleria mellonellalarvae in a time-dependent manner. DepoKP36 did not affect the antibiotic susceptibility ofKlebsiellastrains. The activity of this enzyme was retained in a broad range of pH values (4.0-7.0) and temperatures (up to 45 °C). Consistently, the circular dichroism (CD) spectroscopy revealed a highly stability with melting transition temperature (Tm) = 65 °C. In contrast to other phage tailspike proteins, this enzyme was susceptible to sodium dodecyl sulfate (SDS) denaturation and proteolytic cleavage. The structural studies in solution showed a trimeric arrangement with a high β-sheet content. Our findings identify depoKP36 as a suitable candidate for the development of new treatments forK. pneumoniaeinfections.
authors.affiliation: Institute of Genetics and Microbiology, University of Wroclaw, Przybyszewskiego 63/77, 51-148 Wroclaw, Poland. grazyna.majkowska-skrobek@uwr.edu.pl.
authors.forename: Grażyna
authors.initials: G
authors.lastname: Majkowska-Skrobek
authors.affiliation: Institute of Genetics and Microbiology, University of Wroclaw, Przybyszewskiego 63/77, 51-148 Wroclaw, Poland. agnieszkalatka1989@gmail.com.
authors.forename: Agnieszka
authors.initials: A
authors.lastname: Łątka
authors.affiliation: Institute of Biostructures and Bioimaging, National Research Council, Via Mezzocannone 16, I-80134 Naples, Italy. rita.berisio@unina.it.
authors.forename: Rita
authors.initials: R
authors.lastname: Berisio
authors.affiliation: Institute of Genetics and Microbiology, University of Wroclaw, Przybyszewskiego 63/77, 51-148 Wroclaw, Poland. barbara-boczkowska@wp.pl.
authors.forename: Barbara
authors.initials: B
authors.lastname: Maciejewska
authors.affiliation: Institute of Biostructures and Bioimaging, National Research Council, Via Mezzocannone 16, I-80134 Naples, Italy. squegliaflavia@gmail.com.
authors.forename: Flavia
authors.initials: F
authors.lastname: Squeglia
authors.affiliation: Institute of Biostructures and Bioimaging, National Research Council, Via Mezzocannone 16, I-80134 Naples, Italy. romanom87@hotmail.it.
authors.forename: Maria
authors.initials: M
authors.lastname: Romano
authors.affiliation: Laboratory of Gene Technology, KU Leuven, Kasteelpark Arenberg 21, box 2462, B-3001 Leuven, Belgium. rob.lavigne@kuleuven.be.
authors.forename: Rob
authors.initials: R
authors.lastname: Lavigne
authors.affiliation: Department of Microbiology and Infection Control, Statens Serum Institut, Artillerivej 5, DK-2300S Copenhagen, Denmark. CAS@ssi.dk.|World Health Organization Collaborating Centre for Reference and Research on Escherichia and Klebsiella, Statens Serum Institut, Artillerivej 5, DK-2300S Copenhagen, Denmark. CAS@ssi.dk.
authors.forename: Carsten
authors.initials: C
authors.lastname: Struve
authors.affiliation: Institute of Genetics and Microbiology, University of Wroclaw, Przybyszewskiego 63/77, 51-148 Wroclaw, Poland. zuzanna.drulis-kawa@uwr.edu.pl.
authors.forename: Zuzanna
authors.initials: Z
authors.lastname: Drulis-Kawa
mesh_terms: D000818:Animals; D000900:Anti-Bacterial Agents / Q000737:chemistry / Q000378:metabolism* / Q000627:therapeutic use; D016667:Bacterial Capsules / Q000378:metabolism*; D001435:Bacteriophages / Q000201:enzymology*; D002942:Circular Dichroism; D003001:Cloning, Molecular; D004195:Disease Models, Animal; D004795:Enzyme Stability; D015870:Gene Expression; D006026:Glycoside Hydrolases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism* / Q000627:therapeutic use; D006863:Hydrogen-Ion Concentration; D007710:Klebsiella Infections / Q000382:microbiology / Q000517:prevention & control; D007711:Klebsiella pneumoniae / Q000187:drug effects* / Q000821:virology*; D007915:Lepidoptera; D008970:Molecular Weight; D011487:Protein Conformation / Q000528:radiation effects; D013696:Temperature; D016896:Treatment Outcome
chemical_list: D000900:Anti-Bacterial Agents; D006026:Glycoside Hydrolases
keywords: Klebsiella sp.; bacteriophage; capsule; polysaccharide depolymerase