Activity-Based Protein Profiling Reveals That Cephalosporins Selectively Active on Non-replicating Mycobacterium tuberculosis Bind Multiple Protein Families and Spare Peptidoglycan Transpeptidases.

Landys Lopez Quezada,Jeffrey Aubé,Ben Gold,Julia Roberts,Sae Woong Park,Jean-Emmanuel Hugonnet,Frank J Schoenen,Zainab Edoo,James C Sacchettini,Robert Smith,Quyen Nguyen,Yan Ling,Kelin Li,Michel Arthur,Carl Nathan,Xiaojun Li,Tania J Lupoli

doi:10.3389/fmicb.2020.01248

Abstract

As β-lactams are reconsidered for the treatment of tuberculosis (TB), their targets are assumed to be peptidoglycan transpeptidases, as verified by adduct formation and kinetic inhibition of Mycobacterium tuberculosis (Mtb) transpeptidases by carbapenems active against replicating Mtb. Here, we investigated the targets of recently described cephalosporins that are selectively active against non-replicating (NR) Mtb. NR-active cephalosporins failed to inhibit recombinant Mtb transpeptidases. Accordingly, we used alkyne analogs of NR-active cephalosporins to pull down potential targets through unbiased activity-based protein profiling and identified over 30 protein binders. None was a transpeptidase. Several of the target candidates are plausibly related to Mtb’s survival in an NR state. However, biochemical tests and studies of loss of function mutants did not identify a unique target that accounts for the bactericidal activity of these beta-lactams against NR Mtb. Instead, NR-active cephalosporins appear to kill Mtb by collective action on multiple targets. These results highlight the ability of these β-lactams to target diverse classes of proteins.

Highlights

The chemotherapy of tuberculosis (TB) remains challenging, with an urgent need for shorter, safer treatment whose effectiveness extends to TB resistant to current regimens (Murray et al, 2016)
One strategy involved a search for drugs that can kill Mycobacterium tuberculosis (Mtb) in vitro when it has been made NR by conditions that mimic those found in the host (Bryk et al, 2008; Mak et al, 2012; Grant et al, 2013; Gold and Nathan, 2017)
The cell wall of mycobacteria was considered refractory to inhibition by β-lactams for decades following the demonstration that penicillin was inactive against Mtb (Abraham et al, 1941). β-Lactams are the most highly prescribed antibiotics in modern medicine (Bush and Macielag, 2010) and have utility in diverse infections (Wivagg et al, 2014)

Summary

Introduction

The chemotherapy of tuberculosis (TB) remains challenging, with an urgent need for shorter, safer treatment whose effectiveness extends to TB resistant to current regimens (Murray et al, 2016). Cure of a high proportion of patients with drug-sensitive TB depends on use of multiple antibiotics for at least 6 months. In part, this is thought to reflect the phenotypic antimicrobial. Non-classical Cephalosporin Targets in Mtb resistance that is characteristic of non-replicating (NR) bacterial populations (Nathan, 2012). To this end, several strategies have been proposed to improve the treatment course against NR bacteria (Murray et al, 2016; Gold and Nathan, 2017). In Mtb, as much as 80% of peptidoglycan crosslinks are mediated by Ldts (Lavollay et al, 2008) when cells are in stationary phase compared to 30–40% of the crosslinks in replicating cells (Wietzerbin et al, 1974)

Methods

Results

Conclusion