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

Abstract

Pattern recognition molecules (PRMs) form an important part of innate immunity, where they facilitate the response to infections and damage by 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 complete high-resolution structural information exists about long pentraxins, unlike the short pentraxins, where there is an abundance of both X-ray and cryoelectron microscopy (cryo-EM)-derived structures. This study presents a high-resolution structure of the prototypical long pentraxin, PTX3. Cryo-EM 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 stabilized by an extensive disulfide network. The cryo-EM 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 cryo-EM two-dimensional averages. The resulting hybrid cryo-EM/AlphaFold structure allowed mapping of ligand binding sites, such as C1q and fibroblast growth factor-2, as well as rationalization 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.

Improved therapeutic consistency and efficacy of CD317+ MSCs through stabilizing TSG6 by PTX3

“…Furthermore, TSG6 has been demonstrated as a biomarker to predict the therapeutic efficacy of MSCs in vivo [ 24 ]. PTX3 could bind to TSG6 directly [ 52 , 53 ]. Therefore, we wondered that whether PTX3 could regulate the expression or activity of TSG6.…”

“…Knocking-down PTX3 significantly reduced the immune suppression function of MSCs and their therapeutic effect, indicating a potential marker for improving the therapeutic effects of MSCs [ 78 , 79 ]. Moreover, the PTX3 and TSG6 are coordinated expressed under inflammation stresses [ 80 ], and they two interact with each other [ 52 , 53 , 81 , 82 ]. And TSG6 is the major effector contributing to the therapeutic effects of MSCs in treating IBD with strong immune suppression capabilities [ 1 , 11 , 48 – 51 ].…”

Probing Phosphorylation-Induced Vibrational Couplings in CFTR by 2D IR Spectra Simulations

“…The biophysical mechanisms of the regulatory effect of R domain have remained elusive, largely due to the lack of complete structure information of R domain, especially in the field of theoretical research. The complete structures predicted by AlphaFold (15, 16) provide possibilities for research in this filed. In addition, due to the inherent disorder of the R domain in physiological states, Molecular Dynamics (MD) method based on force field derivation of atomic coordinates can fully demonstrate this disorder process on the time scale, which is a naturalistic method for studying disorder structures.…”

Structural insights into the biological functions of the long pentraxin PTX3

“…Also, a 3D model of human CRP is shown [PDB-ID: 1GNH and ( 8 )] that highlights the symmetric pentameric quaternary structure of this short pentraxin. (B) Orthogonal views of a high-resolution model of the C-terminal pentraxin domains of the human PTX3 based on Cryo-EM that show these domains to fold into octamers with D4 symmetry [PDB-ID: 7ZL1 and ( 9 )]. (C) Schematic drawing of the PTX3 protein with the N-terminal regions (in yellow) forming two parallel tetrameric coiled coils at the opposite sides of the C-terminal core (in red) [based on ( 9 )].…”

“…(B) Orthogonal views of a high-resolution model of the C-terminal pentraxin domains of the human PTX3 based on Cryo-EM that show these domains to fold into octamers with D4 symmetry [PDB-ID: 7ZL1 and ( 9 )]. (C) Schematic drawing of the PTX3 protein with the N-terminal regions (in yellow) forming two parallel tetrameric coiled coils at the opposite sides of the C-terminal core (in red) [based on ( 9 )]. Hinge and intrinsically disordered regions (represented by black lines) bring flexibility to the structure.…”