title: Francisella DnaK inhibits tissue-nonspecific alkaline phosphatase.
abstract: Following pulmonary infection with Francisella tularensis, we observed an unexpected but significant reduction of alkaline phosphatase, an enzyme normally up-regulated following inflammation. However, no reduction was observed in mice infected with a closely related gram-negative pneumonic organism (Klebsiella pneumoniae) suggesting the inhibition may be Francisella-specific. In similar fashion to in vivo observations, addition of Francisella lysate to exogenous alkaline phosphatase (tissue-nonspecific isozyme) was inhibitory. Partial purification and subsequent proteomic analysis indicated the inhibitory factor to be the heat shock protein DnaK. Incubation with increasing amounts of anti-DnaK antibody reduced the inhibitory effect in a dose-dependent manner. Furthermore, DnaK contains an adenosine triphosphate binding domain at its N terminus, and addition of adenosine triphosphate enhances dissociation of DnaK with its target protein, e.g. alkaline phosphatase. Addition of adenosine triphosphate resulted in decreased DnaK co-immunoprecipitated with alkaline phosphatase as well as reduction of Francisella-mediated alkaline phosphatase inhibition further supporting the binding of Francisella DnaK to alkaline phosphatase. Release of DnaK via secretion and/or bacterial cell lysis into the extracellular milieu and inhibition of plasma alkaline phosphatase could promote an orchestrated, inflammatory response advantageous to Francisella.
authors.affiliation: Department of Biology, University of Texas, San Antonio, Texas 78249, USA.
authors.forename: Bernard P
authors.initials: BP
authors.lastname: Arulanandam
authors.forename: Senthilnath Lakshmana
authors.initials: SL
authors.lastname: Chetty
authors.forename: Jieh-Juen
authors.initials: JJ
authors.lastname: Yu
authors.forename: Sean
authors.initials: S
authors.lastname: Leonard
authors.forename: Karl
authors.initials: K
authors.lastname: Klose
authors.forename: Janakiram
authors.initials: J
authors.lastname: Seshu
authors.forename: Andrew
authors.initials: A
authors.lastname: Cap
authors.forename: James J
authors.initials: JJ
authors.lastname: Valdes
authors.forename: James P
authors.initials: JP
authors.lastname: Chambers
mesh_terms: D000255:Adenosine Triphosphate / Q000737:chemistry; D000469:Alkaline Phosphatase / Q000037:antagonists & inhibitors / Q000097:blood*; D000818:Animals; D016470:Bacteremia / Q000382:microbiology*; D058491:Bacterial Load; D001426:Bacterial Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism / Q000502:physiology*; D018834:Chaperonin 60 / Q000378:metabolism; D002848:Chromatography, DEAE-Cellulose; D005260:Female; D005603:Francisella / Q000502:physiology*; D018840:HSP70 Heat-Shock Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism / Q000502:physiology*; D018841:HSP90 Heat-Shock Proteins / Q000378:metabolism; D054884:Host-Pathogen Interactions; D051379:Mice; D008807:Mice, Inbred BALB C; D008970:Molecular Weight; D010446:Peptide Fragments / Q000737:chemistry; D011485:Protein Binding; D021241:Spectrometry, Mass, Electrospray Ionization; D014406:Tularemia / Q000382:microbiology*
chemical_list: D001426:Bacterial Proteins; D018834:Chaperonin 60; D018840:HSP70 Heat-Shock Proteins; D018841:HSP90 Heat-Shock Proteins; D010446:Peptide Fragments; C066720:HtpG protein, bacteria; D000255:Adenosine Triphosphate; C510878:ALPL protein, mouse; D000469:Alkaline Phosphatase