skip to main content


Title: Mechanistic insights into c-di-GMP–dependent control of the biofilm regulator FleQ from Pseudomonas aeruginosa

Bacterial biofilm formation during chronic infections confers increased fitness, antibiotic tolerance, and cytotoxicity. In many pathogens, the transition from a planktonic lifestyle to collaborative, sessile biofilms represents a regulated process orchestrated by the intracellular second-messenger c-di-GMP. A main effector for c-di-GMP signaling in the opportunistic pathogen Pseudomonas aeruginosa is the transcription regulator FleQ. FleQ is a bacterial enhancer-binding protein (bEBP) with a central AAA+ ATPase σ 54-interaction domain, flanked by a C-terminal helix-turn-helix DNA-binding motif and a divergent N-terminal receiver domain. Together with a second ATPase, FleN, FleQ regulates the expression of flagellar and exopolysaccharide biosynthesis genes in response to cellular c-di-GMP. Here we report structural and functional data that reveal an unexpected mode of c-di-GMP recognition that is associated with major conformational rearrangements in FleQ. Crystal structures of FleQ’s AAA+ ATPase domain in its apo-state or bound to ADP or ATP-γ-S show conformations reminiscent of the activated ring-shaped assemblies of other bEBPs. As revealed by the structure of c-di-GMP–complexed FleQ, the second messenger interacts with the AAA+ ATPase domain at a site distinct from the ATP binding pocket. c-di-GMP interaction leads to active site obstruction, hexameric ring destabilization, and discrete quaternary structure transitions. Solution and cell-based studies confirm couplingmore » of the ATPase active site and c-di-GMP binding, as well as the functional significance of crystallographic interprotomer interfaces. Taken together, our data offer unprecedented insight into conserved regulatory mechanisms of gene expression under direct c-di-GMP control via FleQ and FleQ-like bEBPs.« less
 [1] ;  [2] ;  [3] ;  [3] ;  [4] ;  [1]
  1. Univ. of Sao Paulo, Sao Carlos (Brazil). Inst. of Physics of Sao Carlos. Dept. of Physics and Interdisciplinary Science
  2. Pasteur Inst., Paris (France). Dept. of Structural Biology and Chemistry; Cornell Univ., Ithaca, NY (United States). College of Veterinary Medicine. Dept. of Molecular Medicine
  3. Univ. of Washington, Seattle, WA (United States). Dept. of Microbiology
  4. Cornell Univ., Ithaca, NY (United States). College of Veterinary Medicine. Dept. of Molecular Medicine
Publication Date:
Grant/Contract Number:
AC02-06CH11357; DMR-1332208; GM-103485; P41-GM103403; 2009/13238-0; 2011/24168-2; R01-AI097307; R01-GM56665
Published Article
Journal Name:
Proceedings of the National Academy of Sciences of the United States of America
Additional Journal Information:
Journal Volume: 113; Journal Issue: 2; Journal ID: ISSN 0027-8424
National Academy of Sciences, Washington, DC (United States)
Research Org:
Cornell Univ., Ithaca, NY (United States); Univ. of Washington, Seattle, WA (United States); Univ. of Sao Paulo, Sao Carlos (Brazil)
Sponsoring Org:
USDOE Office of Science (SC); National Science Foundation (NSF); National Inst. of Health (NIH) (United States); European Research Council (ERC); São Paulo Research Foundation (Brazil)
Contributing Orgs:
Pasteur Inst., Paris (France)
Country of Publication:
United States
59 BASIC BIOLOGICAL SCIENCES; 60 APPLIED LIFE SCIENCES; enhancer binding protein; flagella; structure; gene expression
OSTI Identifier:
Alternate Identifier(s):
OSTI ID: 1235479