Title
EF-hand protein, EfhP, specifically binds Ca2+ and mediates Ca2+ regulation of virulence in a human pathogen Pseudomonas aeruginosa
Author
Biraj B. Kayastha, Aya Kubo, Jacob Burch-Konda, Rosalie L. Dohmen, Jacee L. McCoy, Rendi R. Rogers, Sergio Mares, Justin Bevere, Annalisa Huckaby, William Witt, Shuxia Peng, Bharat Chaudhary, Smita Mohanty, Mariette Barbier, Gabriel Cook, Junpeng Deng & Marianna A. Patrauchan
Year
2022
Journal
Scientific Reports
Abstract
Calcium (Ca2+) is well known as a second messenger in eukaryotes, where Ca2+ signaling controls life-sustaining cellular processes. Although bacteria produce the components required for Ca2+ signaling, little is known about the mechanisms of bacterial Ca2+ signaling. Previously, we have identified a putative Ca2+-binding protein EfhP (PA4107) with two canonical EF-hand motifs and reported that EfhP mediates Ca2+ regulation of virulence factors production and infectivity in Pseudomonas aeruginosa, a human pathogen causing life-threatening infections. Here, we show that EfhP selectively binds Ca2+ with 13.7 µM affinity, and that mutations at the +X and −Z positions within each or both EF-hand motifs abolished Ca2+ binding. We also show that the hydrophobicity of EfhP increased in a Ca2+-dependent manner, however no such response was detected in the mutated proteins. 15 N-NMR showed Ca2+-dependent chemical shifts in EfhP confirming Ca2+-binding triggered structural rearrangements in the protein. Deletion of efhP impaired P. aeruginosa survival in macrophages and virulence in vivo. Disabling EfhP Ca2+ binding abolished Ca2+ induction of pyocyanin production in vitro. These data confirm that EfhP selectively binds Ca2+, which triggers its structural changes required for the Ca2+ regulation of P. aeruginosa virulence, thus establishing the role of EfhP as a Ca2+ sensor.
Full Article
Instrument
J-810
Keywords
Calcium, EfhP, PA4107, affinity