Phenotypic profiling of mycobacterium tuberculosis espa point mutants reveals that blockage of ESAT-6 and CFP-10 secretion in vitro does not always correlate with attenuation of virulence

Jeffrey M. Chen, Ming Zhang, Jan Rybniker, Laetitia Basterra, Neeraj Dhar, Anna D. Tischler, Florence Pojer, Stewart T. Cole

Research output: Contribution to journalArticlepeer-review

25 Scopus citations

Abstract

The EspA protein of Mycobacterium tuberculosis is essential for the type VII ESX-1 protein secretion apparatus, which delivers the principal virulence factors ESAT-6 and CFP-10. In this study, site-directed mutagenesis of EspA was performed to elucidate its influence on the ESX-1 system. Replacing Trp55 (W55) or Gly57(G57) residues in the putative W-X-G motif of EspA with arginines impaired ESAT-6 and CFP-10 secretion in vitro and attenuated M. tuberculosis. Replacing the Phe50 (F50) and Lys62 (K62) residues, which flank the W-X-G motif, with arginine and alanine, respectively, destabilized EspA, abolished ESAT-6 and CFP-10 secretion in vitro, and attenuated M. tuberculosis. Likewise, replacing the Phe5 (F5) and Lys41 (K41) residues with arginine and alanine, respectively, also destabilized EspA and blocked ESAT-6 and CFP-10 secretion in vitro. However, these two particular mutations did not attenuate M. tuberculosis in cellular models of infection or during acute infection in mice. We have thus identified amino acid residues in EspA that are important for facilitating ESAT-6 and CFP-10 secretion and virulence. However, our data also indicate for the first time that blockage of M. tuberculosis ESAT-6 and CFP-10 secretion in vitro and attenuation are mutually exclusive.

Original languageEnglish (US)
Pages (from-to)5421-5430
Number of pages10
JournalJournal of bacteriology
Volume195
Issue number24
DOIs
StatePublished - Dec 2013

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