An Oligopeptide Transporter of Mycobacterium tuberculosis Regulates Cytokine Release and Apoptosis of Infected Macrophages

Background: The Mycobacterium tuberculosis genome encodes two peptide transporters encoded by Rv3665c-Rv3662c and Rv1280c-Rv1283c. Both belong to the family of ABC transporters containing two nucleotide-binding subunits, two integral membrane proteins and one substrate-binding polypeptide. However, little is known about their functions in M. tuberculosis. Here we report functional characterization of the Rv1280c-Rv1283c-encoded transporter and its substrate-binding polypeptide OppA(MTB). Methodology/Principal Findings: OppA(MTB) was capable of binding the tripeptide glutathione and the nonapeptide bradykinin, indicative of a somewhat broad substrate specificity. Amino acid residues G109, N110, N230, D494 and F496, situated at the interface between domains I and III of OppA, were required for optimal peptide binding. Complementaton of an oppA knockout mutant of M. smegmatis with OppA(MTB) confirmed the role of this transporter in importing glutathione and the importance of the aforesaid amino acid residues in peptide transport. Interestingly, this transporter regulated the ability of M. tuberculosis to lower glutathione levels in infected compared to uninfected macrophages. This ability was partly offset by inactivation of oppD. Concomitantly, inactivation of oppD was associated with lowered levels of methyl glyoxal in infected macrophages and reduced apoptosis-inducing ability of the mutant. The ability to induce the production of the cytokines IL-1 beta, IL-6 and TNF-alpha was also compromised after inactivation of oppD. Conclusions: Taken together, these studies uncover the novel observations that this peptide transporter modulates the innate immune response of macrophages infected with M. tuberculosis

Three-dimensional structure of neurotoxin-1 from Naja naja oxiana venom at 1.9 Å resolution

AbstractNeurotoxin-1 from Naja naja oxiana venom (NTX-1) has been crystallized by vapor diffusion in sitting drops. The crystals have cell dimensions of a = 25.2 Å,b = 75.6 Å, c = 35.9 Å, and are in space group P212121. Three-dimensional data to 1.9 Å have been recorded by a Syntex P21 automatic diffractometer. The atomic structure of the toxin has been determined by molecular replacement using the α-cobratoxin (α-CTX) as the search model. The position of 534 non-hydrogen protein atoms have been determined. The model contains 65 water molecules. Refinement has led to an R-factor of 19.3% at 1.9 Å resolution. The secondary and tertiary structures of NTX-1 have been analyzed and a comparison with structure of the α-CTX has been made

Crystal Structure of the VHS and FYVE Tandem Domains of Hrs, a Protein Involved in Membrane Trafficking and Signal Transduction

AbstractWe have determined the 2 Å X-ray structure of the 219-residue N-terminal VHS and FYVE tandem domain unit of Drosophila Hrs. The unit assumes a pyramidal structure in which the much larger VHS domain (residues 1–153) forms a rectangular base and the FYVE domain occupies the apical end. The VHS domain is comprised of an unusual “superhelix” of eight α helices, and the FYVE domain is mainly built of loops, two double-stranded antiparallel sheets, and a helix stabilized by two tetrahedrally coordinated zinc atoms. The two-domain structure forms an exact 2-fold-related homodimer through antiparallel association of mainly FYVE domains. Dimerization creates two identical pockets designed for binding ligands with multiple negative charges such as citrate or phosphatidylinositol 3-phosphate