PTX3 structure determination using a hybrid cryo-electron microscopy and AlphaFold approach offers insights into ligand binding and complement activation

ABSTRACT: Pattern recognition molecules (PRMs) form an important part of innate immunity and respond to infection and damage via triggering processes such as inflammation. The pentraxin family of soluble PRMs comprises long and short pentraxins, with the former containing unique N-terminal regions unrelated to other proteins or each other. No high-resolution structural information exists about long pentraxins, unlike the short pentraxins where there is an abundance of both X-ray and cryo-electron microscopy (cryoEM) derived structures. This study presents for the first time a high-resolution structure of the prototypical long pentraxin, PTX3. CryoEM yielded a 2.5 Å map of the C-terminal pentraxin domains that revealed a radically different quaternary structure compared to other pentraxins, comprising a glycosylated D4 symmetrical octameric complex stabilised by an extensive disulfide network. The cryoEM map indicated α-helices that extended N-terminal of the pentraxin domains that were not fully resolved. AlphaFold was used to predict the remaining N-terminal structure of the octameric PTX3 complex, revealing two long tetrameric coiled coils with two hinge regions, which was validated using classification of cryoEM 2D averages. The resulting hybrid cryoEM/AlphaFold structure allowed mapping of ligand binding sites, such as C1q and FGF2, as well as rationalisation of previous biochemical data. Given the relevance of PTX3 in conditions ranging from COVID-19 prognosis, cancer progression and female infertility, this structure could be used to inform the understanding and rational design of therapies for these disorders and processes.

INSTRUMENT(S): Orbitrap Fusion Lumos

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Kidney

SUBMITTER: Yassene Mohammed  

LAB HEAD: Paul J. Hensbergen

PROVIDER: PXD034602 | Pride | 2022-10-14

REPOSITORIES: Pride