期刊
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA
卷 117, 期 29, 页码 17211-17220出版社
NATL ACAD SCIENCES
DOI: 10.1073/pnas.2001232117
关键词
c-di-GMP; biofilm; peptide design; protein dynamics; NMR solution structure
资金
- Swiss Commission for Technology and Innovation [18366.1 PFLS-LS]
- Swiss National Science Foundation [31-149927, 31173089, 31-166652, 31-147090]
The bacterial second messenger cyclic diguanylate (c-di-GMP) regulates a wide range of cellular functions from biofilm formation to growth and survival. Targeting a second-messenger network is challenging because the system involves a multitude of components with often overlapping functions. Here, we present a strategy to intercept c-di-GMP signaling pathways by directly targeting the second messenger. For this, we developed a c-di-GMPsequestering peptide (CSP) that was derived from a CheY-like c-di-GMP effector protein. CSP binds c-di-GMP with submicromolar affinity. The elucidation of the CSP center dot c-di-GMP complex structure by NMR identified a linear c-di-GMP-binding motif, in which a self-intercalated c-di-GMP dimer is tightly bound by a network of H bonds and pi-stacking interactions involving arginine and aromatic residues. Structure-based mutagenesis yielded a variant with considerably higher, low-nanomolar affinity, which subsequently was shortened to 19 residues with almost uncompromised affinity. We demonstrate that endogenously expressed CSP intercepts c-diGMP signaling and effectively inhibits biofilm formation in Pseudomonas aeruginosa, the most widely used model for serious biofilm-associated medical implications.
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