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Research Themes Drug discovery

Infectious Diseases: Determining the Essential Structome

SBKB [doi:10.1038/sbkb.2012.141]
Technical Highlight - May 2013
Short description: Twenty-five potential drug targets emerge from the Burkholderia structome through an integrated pipeline.

Binding pocket of isochorismatase, an enzyme identified as essential in B. thailandensis by transposon mutagenesis. The pocket structure provides a template for drug design. Figure courtesy of Wesley Van Voorhis.

The bacterium Burkholderia pseudomallei is the causative agent of melioidosis, a disease endemic to parts of Australia and Southeast Asia. Burkholderia species also cause glanders in animals and pulmonary infections in patients with partially compromised immune systems. The development of new antibiotics is urgently needed, as drug-resistant strains continue to emerge and spread. While structure-based drug design is promising, challenges remain in the search for new antimicrobial agents, such as avoiding the large, non-essential proteome and the difficulties of structure determination.

Overcoming these limitations, Van Voorhis and colleagues (SSCGID) have taken an integrated functional and structural genomics approach to finding potential targets for antibiotics in Burkholderia. First, 406 putative essential genes in the low-virulence B. thailandensis species were identified by saturation-level transposon mutagenesis followed by sequencing. Next, a battery of biological and biochemical criteria narrowed the list down to 315 genes for expression and structure determination.

Typically, the success rate of structure determination from gene candidates is lower than 10%, due to insoluble expression and crystallization difficulties. The authors met this challenge by implementing an “ortholog rescue” in which 387 homologous genes from seven other Burkholderia species were added to the structure-determination pipeline. 450 were expressed in soluble form and 170 crystalized; 68 crystals diffracted with sufficient resolution to meet quality criteria. In total, 88 new Burkholderia protein structures were deposited in the PDB, covering 56 Burkholderia proteins and 49 of the 406 B. thailandensis putative essential genes (a 12.1% gene-to-structure rate).

Interestingly, the underlying rationale for the “ortholog rescue” approach can be verified with the experimental data from seven pairs of ortholog structures solved in the new study. The average overall Cα r.m.s.d. of all pairs was 1.5±0.5Å, indicating that ortholog structures are similar enough to serve as surrogates in drug discovery.

Finally, 25 potential drug targets were identified using three criteria: a lack of close human homologs, membership in an essential metabolic pathway and possession of an essential drug-binding pocket. The authors described five of the most interesting candidates, belonging to synthesis pathways of fatty acids, lipopolysaccharides, siderophores and nucleic acids. As part of the NIAID center's mission, expression clones and proteins were also made available to the infectious disease community for developing new anti-Burkholderia agents.

Wayne Peng

References

  1. L. Baugh et al. Combining functional and structural genomics to sample the essential Burkholderia structome.
    PLoS ONE. 8, e53851 (2013). doi:10.1371/journal.pone.0053851

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