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Structure–function analyses of the bacterial zinc metalloprotease effector protein GtgA uncovers key residues required for deactivating NF-κB

DOI: 10.1074/jbc.RA118.004255 DOI Help

Authors: Elliott Jennings (Imperial College London) , Diego Esposito (Francis Crick Institute) , Katrin Rittinger (The Francis Crick Institute) , Teresa L. M. Thurston (Imperial College London)
Co-authored by industrial partner: No

Type: Journal Paper
Journal: Journal Of Biological Chemistry

State: Published (Approved)
Published: July 2018
Diamond Proposal Number(s): 13775

Open Access Open Access

Abstract: The closely related type III secretion system zinc metalloprotease effector proteins GtgA, GogA, and PipA are translocated into host cells during Salmonella infection. They then cleave nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) transcription factor subunits, dampening activation of the NF-κB signaling pathway and thereby suppressing host immune responses. We demonstrate here that GtgA, GogA, and PipA cleave a subset of NF-κB subunits including p65, RelB, and cRel but not NF-κB1 and NF-κB2, whereas the functionally similar type III secretion system effector NleC of enteropathogenic and enterohemorrhagic Escherichia coli cleaved all five NF-κB subunits. Mutational analysis of NF-κB subunits revealed that a single nonconserved residue in NF-κB1 and NF-κB2 that corresponds to the P1’ residue Arg-41 in p65, prevents cleavage of these subunits by GtgA, GogA, and PipA, explaining the observed substrate specificity of these enzymes. Crystal structures of GtgA in its apo form and in complex with the p65 N-terminal domain explained the importance of the P1’ residue. Furthermore, the pattern of interactions suggested that GtgA recognizes NF-κB subunits by mimicking the shape and negative charge of the DNA phosphate backbone. Moreover, structure-based mutational analysis of GtgA uncovered amino acids that are required for the interaction of GtgA with p65, as well as those that are required for full activity of GtgA in suppressing NF-κB activation. This study therefore provides detailed and critical insight into the mechanism of substrate recognition by this family of proteins important for bacterial virulence.

Journal Keywords: bacterial effectors; GtgA; virulence factor; Salmonella enterica; substrate specificity; type III secretion system (T3SS); NF-kappa B (NF-KB); metalloprotease; bacterial pathogenesis

Diamond Keywords: Bacteria; Enzymes

Subject Areas: Biology and Bio-materials


Instruments: I04-Macromolecular Crystallography , I24-Microfocus Macromolecular Crystallography

Added On: 09/08/2018 11:12

Documents:
PIIS0021925820353059.pdf

Discipline Tags:

Pathogens Infectious Diseases Health & Wellbeing Structural biology Life Sciences & Biotech

Technical Tags:

Diffraction Macromolecular Crystallography (MX)