title: The PE_PGRS Proteins of Mycobacterium tuberculosis Are Ca(2+) Binding Mediators of Host-Pathogen Interaction.
abstract: The phenomenal success of Mycobacterium tuberculosis (M.tb) as a pathogen is primarily based on its ability to modulate host immune responses. The genome of M.tb encodes multiple immunomodulatory proteins, including several members of the multigenic PE_PPE family of which the PE_PGRS proteins are a subset. Curiously, 56 of the 61 PE_PGRS proteins contain multiple copies of the glycine-rich sequence motif GGXGXD/NXUX, a nonapeptide sequence predicted to bind Ca(2+), but the functional significance of these motifs remains a mystery. Here we provide evidence via isothermal titration calorimetry, (45)Ca blotting, fluorescence, and circular dichroism spectroscopy that Ca(2+) binds to the PE_PGRS proteins, PE_PGRS33 (Rv1818c) (10 motifs) and PE_PGRS61 (Rv3653) (one motif). Ca(2+) was observed not to bind to PE_PGRS8 (Rv0742), which lacks nonapeptide motifs. Using recombinant Mycobacterium smegmatis strains expressing Rv1818c and Rv3653 and the THP-1 macrophage model of infection, we show that the two proteins mediate Ca(2+)-dependent upregulation of the anti-inflammatory cytokine IL-10, events critical to the pathogenesis of M.tb. Both Rv1818c and Rv3653 interact with TLR2 in a Ca(2+)-dependent manner, providing a novel mechanistic basis for their immunomodulatory effects. Mutations in the nonapeptide motif of Rv3653 led to compromised Ca(2+) binding, validating the functional criticality of this motif. This study demonstrates for the first time not only their Ca(2+) binding properties but also an essential role for Ca(2+) in the functioning of the M.tb PE_PGRS proteins, opening up the possibility of developing novel anti-tuberculosis therapeutics that inhibit Ca(2+)-PE_PGRS binding.
authors.affiliation: CSIR-Centre for Cellular and Molecular Biology , Uppal Road, Hyderabad 500007, India.
authors.forename: Veena C
authors.initials: VC
authors.lastname: Yeruva
authors.affiliation: CSIR-Centre for Cellular and Molecular Biology , Uppal Road, Hyderabad 500007, India.
authors.forename: Apoorva
authors.initials: A
authors.lastname: Kulkarni
authors.affiliation: CSIR-Centre for Cellular and Molecular Biology , Uppal Road, Hyderabad 500007, India.
authors.forename: Radhika
authors.initials: R
authors.lastname: Khandelwal
authors.affiliation: CSIR-Centre for Cellular and Molecular Biology , Uppal Road, Hyderabad 500007, India.
authors.forename: Yogendra
authors.initials: Y
authors.lastname: Sharma
authors.affiliation: CSIR-Centre for Cellular and Molecular Biology , Uppal Road, Hyderabad 500007, India.
authors.forename: Tirumalai R
authors.initials: TR
authors.lastname: Raghunand
mesh_terms: D001426:Bacterial Proteins / Q000737:chemistry*; D002118:Calcium / Q000737:chemistry*; D002942:Circular Dichroism; D054884:Host-Pathogen Interactions; D009169:Mycobacterium tuberculosis / Q000737:chemistry*; D013050:Spectrometry, Fluorescence
chemical_list: D001426:Bacterial Proteins; D002118:Calcium