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Fluorinated trehalose analogues for cell surface engineering and imaging of Mycobacterium tuberculosis.

ABSTRACT: The sensitive, rapid and accurate diagnosis of Mycobacterium tuberculosis (Mtb) infection is a central challenge in controlling the global tuberculosis (TB) pandemic. Yet the detection of mycobacteria is often made difficult by the low sensitivity of current diagnostic tools, with over 3.6 million TB cases missed each year. To overcome these limitations there is an urgent need for next-generation TB diagnostic technologies. Here we report the use of a discrete panel of native ¹⁹F-trehalose (F-Tre) analogues to label and directly visualise Mtb by exploiting the uptake of fluorine-modified trehalose analogues via the mycobacterial trehalose LpqY-SugABC ATP-binding cassette (ABC) importer. We discovered the extent of modified F-Tre uptake correlates with LpqY substrate recognition and characterisation of the interacting sites by saturation transfer difference NMR coupled with molecular dynamics provides a unique glimpse into the molecular basis of fluorine-modified trehalose import in Mtb. Lipid profiling demonstrated that F-Tre analogues modified at positions 2, 3 and 6 are incorporated into mycobacterial cell-surface trehalose-containing glycolipids. This rapid one-step labelling approach facilitates the direct visualisation of F-Tre-labelled Mtb by Focused Ion Beam (FIB) Secondary Ion Mass Spectrometry (SIMS), enabling detection of the Mtb pathogen. Collectively, our findings highlight that F-Tre analogues have potential as tools to probe and unravel Mtb biology and can be exploited to detect and image TB.

SUBMITTER: Guy CS

PROVIDER: S-EPMC11317875 | biostudies-literature | 2024 Aug

REPOSITORIES: biostudies-literature

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Fluorinated trehalose analogues for cell surface engineering and imaging of Mycobacterium tuberculosis.

Guy Collette S CS Gott James A JA Ramírez-Cárdenas Jonathan J de Wolf Christopher C Furze Christopher M CM West Geoff G Muñoz-García Juan C JC Angulo Jesus J Fullam Elizabeth E

Chemical science 20240812

The sensitive, rapid and accurate diagnosis of Mycobacterium tuberculosis (Mtb) infection is a central challenge in controlling the global tuberculosis (TB) pandemic. Yet the detection of mycobacteria is often made difficult by the low sensitivity of current diagnostic tools, with over 3.6 million TB cases missed each year. To overcome these limitations there is an urgent need for next-generation TB diagnostic technologies. Here we report the use of a discrete panel of native ...[more]

PMID: 39144457

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Project description:Mycobacteria and related organisms in the Corynebacterineae suborder are characterized by a distinctive outer membrane referred to as the mycomembrane. Biosynthesis of the mycomembrane occurs through an essential process called mycoloylation, which involves antigen 85 (Ag85)-catalyzed transfer of mycolic acids from the mycoloyl donor trehalose monomycolate (TMM) to acceptor carbohydrates and, in some organisms, proteins. We recently described an alkyne-modified TMM analogue (O-AlkTMM-C7) which, in conjunction with click chemistry, acted as a chemical reporter for mycoloylation in intact cells and allowed metabolic labeling of mycoloylated components of the mycomembrane. Here, we describe the synthesis and evaluation of a toolbox of TMM-based reporters bearing alkyne, azide, trans-cyclooctene, and fluorescent tags. These compounds gave further insight into the substrate tolerance of mycoloyltransferases (e.g., Ag85s) in a cellular context and they provide significantly expanded experimental versatility by allowing one- or two-step cell labeling, live cell labeling, and rapid cell labeling via tetrazine ligation. Such capabilities will facilitate research on mycomembrane composition, biosynthesis, and dynamics. Moreover, because TMM is exclusively metabolized by Corynebacterineae, the described probes may be valuable for the specific detection and cell-surface engineering of Mycobacterium tuberculosis and related pathogens. We also performed experiments to establish the dependence of probe incorporation on mycoloyltransferase activity, results from which suggested that cellular labeling is a function not only of metabolic incorporation (and likely removal) pathway(s), but also accessibility across the envelope. Thus, whole-cell labeling experiments with TMM reporters should be carefully designed and interpreted when envelope permeability may be compromised. On the other hand, this property of TMM reporters can potentially be exploited as a convenient way to probe changes in envelope integrity and permeability, facilitating drug development studies.

Dataset Information

Fluorinated trehalose analogues for cell surface engineering and imaging of Mycobacterium tuberculosis.

Publications

Fluorinated trehalose analogues for cell surface engineering and imaging of <i>Mycobacterium tuberculosis</i>.

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