9R3Y

Solution NMR structure of N-WASP GBD in complex with EspFu R5

Summary for 9R3Y

• Entry DOI: 10.2210/pdb9r3y/pdb
• Related: 2LNH
• NMR Information: BMRB: 34992
• Descriptor: Actin nucleation-promoting factor WASL, Secreted effector protein EspF(U) (2 entities in total)
• Functional Keywords: complex, signaling protein
• Biological source: Homo sapiens (human); More
• Total number of polymer chains: 2
• Total formula weight: 13605.20

Authors

Tossavainen, H.,Permi, P. (deposition date: 2025-05-06, release date: 2025-10-22)

Primary citation

Tossavainen, H.,Karjalainen, M.,Antenucci, L.,Hellman, M.,Permi, P.
Intrinsically disordered enteropathogenic E. coli EspF exploits motif mimicry in high-affinity binding to neural Wiskott-Aldrich syndrome protein and sorting nexin 9.
Int.J.Biol.Macromol., 330:148227-148227, 2025

PubMed Abstract: EspF is an enteropathogenic Escherichia coli (EPEC) effector protein that interferes with intestinal epithelial cell signaling by binding to the Src homology 3 (SH3) domain of sorting nexin 9 (SNX9) and the GTPase-binding domain (GBD) of neural Wiskott-Aldrich syndrome protein (N-WASP) with its C-terminal proline-rich repeats. To understand the molecular basis of these interactions, we characterized the structure, dynamics, and binding thermodynamics of EspF and its target protein domain complexes. We also elaborated on our previous study on EspF, a homologous effector in enterohemorrhagic E. coli (EHEC), and compared the two effectors. We show that EspF is intrinsically disordered but that NMR chemical shifts expose the pre-structured polyproline II (PPII) helical SH3- and helical GBD-binding motifs. These motifs mimic their cellular counterparts but are fine-tuned to prevail in competitive binding. Factors behind EspF's higher affinity for GBD relative to the cellular ligand are key residue mutations and a C-terminally elongated polar interaction interface. The latter compensates for the lack of an "extended arm", the critical substitution promoting high affinity for GBD in EspF. With this advantage, EspF outcompetes the autoinhibitory N-WASP C-helix and stimulates actin polymerization. EspF binds SNX9 SH3 with an extended binding interface, residues N-terminal to the RxAPxxP core motif being essential to strong binding. We define the SNX9 SH3-binding epitope as ϕxPxRxAPxxP and propose to re-delineate the EPEC EspF repeat boundaries accordingly. Furthermore, a characteristic C secondary chemical shift pattern is recognized as a fingerprint of polyproline II (PPII) helical conformation in the SH3 binding epitope.

PubMed: 41075884
DOI: 10.1016/j.ijbiomac.2025.148227

Experimental method

SOLUTION NMR