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The beast and its Achilles heel:
A novel target to fight multi-resistant pathogenic bacteria
Outlook
Worldwide, the number of antibiotic resistant bacteria is increasing, while the development of new antibiotics is still stumbling. Therefore, we intended to build a reporter system to screen for antibiotics directed against cyclic di-AMP, an essential signaling nucleotide found in many Gram-positive bacteria.
We applied the Gram-negative bacterium E. coli , which has no endogenous c-di-AMP, as a carrier organism. We used several existing Biobricks and recently discovered c-di-AMP sensors from Bacillus subtilis or Mycobacterium smegmatis, We managed to construct two in vivo screening systems based on different principals. Though these systems still need improvement, they were active and partially functional in E. coli. Thus, we plan to optimize these systems for further development. Once these systems are fully functional, one could provide E. coli with c-di-AMP by co-expressing its biosynthesis enzyme, a diadenylate cyclase(Fig. 1). Such an in vivo screening system could be used to screen effectively for novel antibiotics.
As c-di-AMP homeostasis plays a vital role in many Gram-positive bacteria, its biosynthesis enzyme, diadenylate cyclase (DAC), becomes a very promising target for novel antibacterial substances. We worked on the characterization of one DAC from Listeria monocytogenes. We were able to obtain the crystal structure with the help of the department of Structural Biology and determined a 3D-structure out of it. With this structure known, in silico screenings for novel antibiotics which could disturb this enzyme could start. The purified DAC could also be employed for in vitro screenings.
Fig. 1. For in vivo screening for new antibiotics, the DAC could be cloned into an E. coli cell harboring a reporter system. The crystal structure of the DAC could be used for in silico screening. The purified enzyme can be used for in vitro screening.
