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J Biol Chem. 2017 Oct 6;292(40):16677-16687. doi: 10.1074/jbc.M117.802231. Epub 2017 Aug 25.

Single-domain antibodies pinpoint potential targets within Shigella invasion plasmid antigen D of the needle tip complex for inhibition of type III secretion.

Author information

1
From the Department of Pharmaceutical Chemistry, University of Kansas, Lawrence, Kansas 66047.
2
Department of Infectious Diseases and Global Health, Tufts Clinical and Translational Science Institute, North Grafton, Massachusetts 02111.
3
Protein Structure Laboratory, Del Shankel Structural Biology Center, University of Kansas, Lawrence, Kansas 66047, and.
4
IMCA-CAT, Hauptman-Woodward Medical Research Institute, Argonne, Illinois 60439.
5
From the Department of Pharmaceutical Chemistry, University of Kansas, Lawrence, Kansas 66047, wendy.picking@ku.edu.

Abstract

Numerous Gram-negative pathogens infect eukaryotes and use the type III secretion system (T3SS) to deliver effector proteins into host cells. One important T3SS feature is an extracellular needle with an associated tip complex responsible for assembly of a pore-forming translocon in the host cell membrane. Shigella spp. cause shigellosis, also called bacillary dysentery, and invade colonic epithelial cells via the T3SS. The tip complex of Shigella flexneri contains invasion plasmid antigen D (IpaD), which initially regulates secretion and provides a physical platform for the translocon pore. The tip complex represents a promising therapeutic target for many important T3SS-containing pathogens. Here, in an effort to further elucidate its function, we created a panel of single-VH domain antibodies (VHHs) that recognize distinct epitopes within IpaD. These VHHs recognized the in situ tip complex and modulated the infectious properties of Shigella Moreover, structural elucidation of several IpaD-VHH complexes provided critical insights into tip complex formation and function. Of note, one VHH heterodimer could reduce Shigella hemolytic activity by >80%. Our observations along with previous findings support the hypothesis that the hydrophobic translocator (IpaB in Shigella) likely binds to a region within the tip protein that is structurally conserved across all T3SS-possessing pathogens, suggesting potential therapeutic avenues for managing infections by these pathogens.

KEYWORDS:

IpaD; Shigella; crystal structure; microbial pathogenesis; single-domain antibody (sdAb, nanobody); type III secretion system (T3SS); virulence factor

PMID:
28842484
PMCID:
PMC5633129
DOI:
10.1074/jbc.M117.802231
[Indexed for MEDLINE]
Free PMC Article

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