9ZDM

RhsF/RhsFi complex from Chromobacterium violaceum

Summary for 9ZDM

• Entry DOI: 10.2210/pdb9zdm/pdb
• Descriptor: RhsFi, Probable Rhs-family protein (3 entities in total)
• Functional Keywords: antibacterial toxin, toxin-immunity complex, adp-ribosyltransferase, antimicrobial protein
• Biological source: Chromobacterium violaceum; More
• Total number of polymer chains: 4
• Total formula weight: 51904.38

Authors

Sgro, G.G.,Alves, J.A.,Pankov, G.,Coulthurst, S.J.,da Silva Neto, J.F. (deposition date: 2025-11-26, release date: 2026-06-17, Last modification date: 2026-07-15)

Primary citation

Alves, J.A.,Pankov, G.,Frey, A.M.,Trost, M.,Sgro, G.G.,Coulthurst, S.J.,da Silva Neto, J.F.
Global identification of Chromobacterium violaceum T6SS effectors reveals an Rhs antibacterial toxin featuring FIX and ADP-ribosyltransferase domains.
J.Biol.Chem., 302:113216-113216, 2026

PubMed Abstract: Bacteria coexist in polymicrobial communities where they engage in complex interactions, including interbacterial antagonism. The environmental bacterial pathogen Chromobacterium violaceum possesses an active type VI secretion system (T6SS), which relies mainly on VgrG3 for its activity and role in interbacterial competition. However, the arsenal of toxic effectors delivered by this T6SS remains unknown. Here, we identify the repertoire of C. violaceum T6SS effectors and characterize a novel antibacterial Rhs-family effector, RhsF (Rhs with a FIX domain), and its cognate immunity protein, RhsFi. Using mass spectrometry analyses of secreted proteins and proteins co-immunoprecipitated with VgrG3, we identified six novel effector candidates, namely four phospholipases, a protein of unknown function, and the previously-uncharacterized Rhs protein, RhsF (CV_1431). RhsF contains an N-terminal FIX domain and can intoxicate susceptible bacteria in a T6SS-dependent manner. The action of the C-terminal toxin domain of RhsF (RhsF-CT) is prevented by RhsFi (CV_1430), confirming that RhsF-RhsFi comprises an effector-immunity pair. The structure of the RhsF-CT/RhsFi complex determined by X-ray crystallography (1.85 Å resolution) revealed that RhsF-CT shares structural similarity with ADP-ribosyltransferase toxins and that RhsFi inhibits toxicity via direct occlusion of the RhsF-CT catalytic site. Functional assays indicated that RhsF-CT ADP-ribosylates RNA in vitro and that RhsF toxicity requires a catalytic triad composed of R1403, Y1456, and E1497 residues. Overall, our findings reveal effectors secreted by the T6SS of C. violaceum, establish RhsF as a potent antibacterial toxin, and confirm T6SS-dependent delivery of a FIX-containing Rhs protein, expanding the known repertoire of bacterial arms involved in microbial competition.

PubMed: 42229804
DOI: 10.1016/j.jbc.2026.113216

Experimental method

X-RAY DIFFRACTION (1.85 Å)