PSI Structural Biology Knowledgebase

PSI | Structural Biology Knowledgebase
Header Icons

Related Articles
Protein Folding and Misfolding: It's the Journey, Not the Destination
March 2015
CCR5 and HIV Infection
January 2015
HIV/AIDS: Pre-fusion Env Exposed
January 2015
HIV/AIDS: Slide to Enter
January 2015
Updating ModBase
January 2015
Power in Numbers
August 2014
Quorum Sensing: A Groovy New Component
August 2014
Bacterial CDI Toxins
June 2014
Immunity: One Antibody to Rule Them All
June 2014
Virology: A Bat Influenza Hemagglutinin
March 2014
Virology: Making Sensitive Magic
March 2014
Virology: Visualizing Cyanophage Assembly
March 2014
Virology: Zeroing in on HBV Egress
March 2014
March 2014
Cas4 Nuclease and Bacterial Immunity
February 2014
Microbial Pathogenesis: A GNAT from Pseudomonas
February 2014
Microbial Pathogenesis: Targeting Drug Resistance in Mycobacterium tuberculosis
February 2014
Microbiome: The Dynamics of Infection
September 2013
Membrane Proteome: A Funnel-like Viroporin
August 2013
Infectious Diseases: A Pathogen Ubiquitin Ligase
May 2013
Infectious Diseases: A Shared Syringe
May 2013
Infectious Diseases: Determining the Essential Structome
May 2013
Infectious Diseases: Targeting Meningitis
May 2013
NDM-1 and Antibiotics
May 2013
Bacterial Hemophores
January 2013
Microbial Pathogenesis: Computational Epitope Prediction
January 2013
Microbial Pathogenesis: Influenza Inhibitor Screen
January 2013
Microbial Pathogenesis: Measles Virus Attachment
January 2013
Microbial Pathogenesis: NEAT Iron
January 2013
Membrane Proteome: Sphingolipid Synthesis Selectivity
December 2012
A signal sensing switch
September 2012
Gauging needle structure
July 2012
Anthrax Stealth Siderophores
June 2012
A Pseudomonas L-serine dehydrogenase
May 2012
Pilus Assembly Protein TadZ
April 2012
Making Lipopolysaccharide
January 2012
Superbugs and Antibiotic Resistance
December 2011
A change to resistance
November 2011
An effective and cooperative dimer
November 2011
The Perils of Protein Secretion
November 2011
Bacterial Armor
October 2011
Breaking down the defenses
September 2011
Moving some metal
August 2011
Capsid assembly in motion
April 2011
Know thy enemy … structurally
October 2010
Treating sleeping sickness
May 2010
Bacterial spore kinase
April 2010
Hemolysin BL
January 2010
Unusual cell division
October 2009
Anthrax evasion tactics
September 2009
Toxin-antitoxin VapBC-5
September 2009
Antibiotic target
August 2009
July 2009
Tackling influenza
June 2009
You look familiar: the Type VI secretion system
June 2009
Unique SARS
April 2009
Anthrax stealth molecule
March 2009
A new class of bacterial E3 ubiquitination enzymes
January 2009
Antiviral evasion
October 2008
SARS connections
September 2008
SARS Coronavirus Nonstructural Protein 1
June 2008

Research Themes Infectious diseases

Microbial Pathogenesis: Targeting Drug Resistance in Mycobacterium tuberculosis

SBKB [doi:10.1038/sbkb.2012.184]
Featured Article - February 2014
Short description: Whole-cell screening for antituberculosis compounds combined with identification of resistance-linked mutations provide novel scaffolds for drug development.

Resistance mutations to compound 4 in the M. tuberculosis aspartyl-tRNA synthetase mapped onto the crystal structure of the AspRS (tRNA:synthetase) complex from T. thermophilus (PDB 1EFW). The resistance mutations map to the dimer interface. Figure courtesy of James Sacchettini.

As the development of drug resistance continues to be a threat to public health, new methods are sought for the rapid identification of novel targets. This is particularly the case for tuberculosis, where the prevalence of drug-resistant strains is rising rapidly. Current antibiotics target a limited number of cellular processes, and efforts in the past two decades to develop new drugs based on pre-specified targets have yielded limited returns.

To identify new drugs effective against the pathogen Mycobacterium tuberculosis, Sacchettini and colleagues (PSI MTBI) have developed a workflow that avoids target bias by combining high-throughput, whole-cell screening with whole- genome sequencing of resistant strains. In addition to eliminating target bias, this platform allows for the rapid identification of a specific target of the lead compound, an essential step for subsequent medicinal chemistry. First, a library of compounds was screened for general inhibition of cell growth, followed by in vitro selection of mutations that confer resistance. To pinpoint the gene product responsible for drug sensitivity, polymorphisms potentially responsible for selected resistance are identified through deep sequencing and confirmed as the basis for resistance by reintroducing single mutations in a clean genetic background, via a phage recombinase.

To demonstrate the utility of this protocol, eight compounds with antitubercular activity were selected from a number of previous whole-cell screens. Several resistant strains were then isolated and sequenced, with the responsible mutations identified. In four cases, the genes associated with the resistant mutations were known to be essential, and thus likely to be the direct targets of the drugs: an ESX-3 type VII secretion system component (EccB3), the aspartyl-tRNA synthetase (AspS), membrane transporter MmpL3 and a polyketide synthase (Pks13). In all but one case, re-engineering the isolated mutations into a wild-type background conferred resistance to the original drug, confirming that the targets had been correctly identified. While MmpL3 had previously been shown to be the target of several small molecules, EccB3, AspS and Pks13 are not targeted by currently available drugs and therefore represent novel target leads. In the case of AspS and Pks13, structural homology modeling also suggested mechanisms of action.

In addition to providing an efficient approach to the identification of novel drug targets in any microorganism, the described protocol can also be used to rapidly build on the data generated by previous whole-cell screens.

Stéphane Larochelle


  1. T.R. Ioerger et al. Identification of new drug targets and resistance mechanisms in Mycobacterium tuberculosis.
    PLoS One. 8, e75245 (2013). doi:10.1371/journal.pone.0075245

Structural Biology Knowledgebase ISSN: 1758-1338
Funded by a grant from the National Institute of General Medical Sciences of the National Institutes of Health