title: Purine biosynthesis is the bottleneck in trimethoprim-treated Bacillus subtilis.
abstract: Trimethoprim is a folate biosynthesis inhibitor. Tetrahydrofolates are essential for the transfer of C1units in several biochemical pathways including purine, thymine, methionine, and glycine biosynthesis. This study addressed the effects of folate biosynthesis inhibition on bacterial physiology.
  
  
  
  Two complementary proteomic approaches were employed to analyze the response of Bacillus subtilis to trimethoprim. Acute changes in protein synthesis rates were monitored by radioactive pulse labeling of newly synthesized proteins and subsequent 2DE analysis. Changes in protein levels were detected using gel-free quantitative MS.
  
  
  
  Proteins involved in purine and histidine biosynthesis, the σB-dependent general stress response, and sporulation were upregulated. Most prominently, the PurR-regulon required for de novo purine biosynthesis was derepressed indicating purine depletion. The general stress response was activated energy dependently and in a subpopulation of treated cultures an early onset of sporulation was observed, most likely triggered by low guanosine triphosphate levels. Supplementation of adenosine triphosphate, adenosine, and guanosine to the medium substantially decreased antibacterial activity, showing that purine depletion becomes the bottleneck in trimethoprim-treated B. subtilis.
  
  
  
  The frequently prescribed antibiotic trimethoprim causes purine depletion in B. subtilis, which can be complemented by supplementing purines to the medium.
authors.affiliation: Ruhr-Universität Bochum, Applied Microbiology, Bochum, Germany.
authors.forename: Jennifer Janina
authors.initials: JJ
authors.lastname: Stepanek
authors.affiliation: Ruhr-Universität Bochum, Applied Microbiology, Bochum, Germany.
authors.forename: Sina
authors.initials: S
authors.lastname: Schäkermann
authors.affiliation: Ruhr-Universität Bochum, Applied Microbiology, Bochum, Germany.
authors.forename: Michaela
authors.initials: M
authors.lastname: Wenzel
authors.affiliation: Ruhr-Universität Bochum, Applied Microbiology, Bochum, Germany.
authors.forename: Pascal
authors.initials: P
authors.lastname: Prochnow
authors.affiliation: Ruhr-Universität Bochum, Applied Microbiology, Bochum, Germany. julia.bandow@rub.de.
authors.forename: Julia Elisabeth
authors.initials: JE
authors.lastname: Bandow
mesh_terms: D000241:Adenosine / Q000494:pharmacology; D000255:Adenosine Triphosphate / Q000494:pharmacology; D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D001412:Bacillus subtilis / Q000187:drug effects* / Q000378:metabolism* / Q000502:physiology; D001426:Bacterial Proteins / Q000378:metabolism; D004734:Energy Metabolism / Q000187:drug effects; D006151:Guanosine / Q000494:pharmacology; D040901:Proteomics; D011687:Purines / Q000096:biosynthesis*; D013171:Spores, Bacterial / Q000187:drug effects / Q000502:physiology; D013312:Stress, Physiological / Q000187:drug effects; D014295:Trimethoprim / Q000494:pharmacology*
chemical_list: D000900:Anti-Bacterial Agents; D001426:Bacterial Proteins; D011687:Purines; D006151:Guanosine; D000255:Adenosine Triphosphate; D014295:Trimethoprim; D000241:Adenosine
keywords: Antibiotic; Sporulation; Trimethoprim; σB-dependent general stress response