Browsing by Author "Lithgow, T"

Now showing 1 - 3 of 3
  • Thumbnail Image
    Item
    Arginine catabolism is essential to polymyxin dependence in Acinetobacter baumannii
    (Elsevier, 2024-07) Han, ML; Alsaadi, Y; Zhao, JX; Zhu, Y; Lu, J; Jiang, X; Ma, W; Patil, NA; Dunstan, RA; Le Brun, AP; Wickremasinghe, H; Hu, X; Wu, Y; Yu, HH; Wang, J; Barlow, CK; Bergen, PJ; Shen, HH; Lithgow, T; Creek, DJ; Velkov, T; Li, J
    Polymyxins are often the only effective antibiotics against the "Critical" pathogen Acinetobacter baumannii. Worryingly, highly polymyxin-resistant A. baumannii displaying dependence on polymyxins has emerged in the clinic, leading to diagnosis and treatment failures. Here, we report that arginine metabolism is essential for polymyxin-dependent A. baumannii. Specifically, the arginine degradation pathway was significantly altered in polymyxin-dependent strains compared to wild-type strains, with critical metabolites (e.g., L-arginine and L-glutamate) severely depleted and expression of the astABCDE operon significantly increased. Supplementation of arginine increased bacterial metabolic activity and suppressed polymyxin dependence. Deletion of astA, the first gene in the arginine degradation pathway, decreased phosphatidylglycerol and increased phosphatidylethanolamine levels in the outer membrane, thereby reducing the interaction with polymyxins. This study elucidates the molecular mechanism by which arginine metabolism impacts polymyxin dependence in A. baumannii, underscoring its critical role in improving diagnosis and treatment of life-threatening infections caused by "undetectable" polymyxin-dependent A. baumannii. ª 2024 The Author(s). Published by Elsevier Inc. This is an open access article under the CC BY-NC licence
  • No Thumbnail Available
    Item
    Domain organization of the monomeric form of the Tom70 mitochondrial import receptor
    (Elsevier, 2009-05-22) Mills, RD; Trewhella, J; Qiu, TW; Welte, T; Ryan, TM; Hanley, TL; Knott, RB; Lithgow, T; Mulhern, TD
    Tom70 is a mitochondrial protein import receptor composed of 11 tetratricopeptide repeats (TPRs). The first three TPRs form an N-terminal domain that recruits heat shock protein family chaperones, while the eight C-terminal TPRs form a domain that receives, from the bound chaperone, mitochondrial precursor proteins destined for import. Analytical Ultracentrifugation and solution small-angle X-ray scattering (SAXS) analysis characterized Tom70 as an elongated monomer. A model for the Tom70 monomer was proposed based on the alternate interpretation of the domain pairings observed in the crystal structure of the Tom70 dimer and refined against the SAXS data. In this "open" model of the Tom70 monomer, the chaperone- and precursor-binding sites are exposed and lay side by side oil one face of the molecule. Fluorescence anisotropy measurements indicated that monomeric Tom70 can bind both chaperone and precursor peptides and that chaperone peptide binding does not alter the affinity of Tom70 for the precursor peptide. SAXS Was unable to detect any shape change in Tom70 upon chaperone binding. However, molecular modeling indicated that chaperone binding is incompatible with Tom70 dimer formation. It is proposed that the Tom70 monomer is the functional unit mediating initial chaperone docking and precursor recognition. © 2009, Elsevier Ltd.
  • No Thumbnail Available
    Item
    Reconstitution of a nanomachine driving the assembly of proteins into bacterial outer membranes
    (Macmillan Publishers Limited., 2014-10-24) Shen, HH; Leyton, DL; Shiota, T; Belousoff, MJ; Noinaj, N; Lu, J; Holt, SA; Tan, K; Selkrig, J; Webb, CT; Buchanan, SK; Martin, LL; Lithgow, T
    In biological membranes, various protein secretion devices function as nanomachines, and measuring the internal movements of their component parts is a major technological challenge. The translocation and assembly module (TAM) is a nanomachine required for virulence of bacterial pathogens. We have reconstituted a membrane containing the TAM onto a gold surface for characterization by quartz crystal microbalance with dissipation (QCM-D) and magnetic contrast neutron reflectrometry (MCNR). The MCNR studies provided structural resolution down to 1 Å, enabling accurate measurement of protein domains projecting from the membrane layer. Here we show that dynamic movements within the TamA component of the TAM are initiated in the presence of a substrate protein, Ag43, and that these movements recapitulate an initial stage in membrane protein assembly. The reconstituted system provides a powerful new means to study molecular movements in biological membranes, and the technology is widely applicable to studying the dynamics of diverse cellular nanomachines. © Macmillan Publishers Limited