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Minicollagen cysteine-rich domains encode distinct modes of polymerization to form stable nematocyst capsules.

Tursch A, Mercadante D, Tennigkeit J, Gräter F, Özbek S - Sci Rep (2016)

Bottom Line: Our combined experimental and computational analyses reveal the cysteines in the C-CRD fold to exhibit a higher structural propensity for disulfide bonding and a faster kinetics of polymerization.During nematocyst maturation, the highly reactive C-CRD is instrumental in efficient cross-linking of minicollagens to form pressure resistant capsules.The higher ratio of C-CRD folding types evidenced in the medusozoan lineage might have fostered the evolution of novel, predatory nematocyst types in cnidarians with a free-swimming medusa stage.

View Article: PubMed Central - PubMed

Affiliation: University of Heidelberg, Centre for Organismal Studies, Department of Molecular Evolution and Genomics, Im Neuenheimer Feld 329, 69120 Heidelberg, Germany.

ABSTRACT
The stinging capsules of cnidarians, nematocysts, function as harpoon-like organelles with unusual biomechanical properties. The nanosecond discharge of the nematocyst requires a dense protein network of the capsule structure withstanding an internal pressure of up to 150 bar. Main components of the capsule are short collagens, so-called minicollagens, that form extended polymers by disulfide reshuffling of their cysteine-rich domains (CRDs). Although CRDs have identical cysteine patterns, they exhibit different structures and disulfide connectivity at minicollagen N and C-termini. We show that the structurally divergent CRDs have different cross-linking potentials in vitro and in vivo. While the C-CRD can participate in several simultaneous intermolecular disulfides and functions as a cystine knot after minicollagen synthesis, the N-CRD is monovalent. Our combined experimental and computational analyses reveal the cysteines in the C-CRD fold to exhibit a higher structural propensity for disulfide bonding and a faster kinetics of polymerization. During nematocyst maturation, the highly reactive C-CRD is instrumental in efficient cross-linking of minicollagens to form pressure resistant capsules. The higher ratio of C-CRD folding types evidenced in the medusozoan lineage might have fostered the evolution of novel, predatory nematocyst types in cnidarians with a free-swimming medusa stage.

No MeSH data available.


Related in: MedlinePlus

Propensity of intermolecular disulfide formation revealed by molecular docking of N-CRD and C-CRD domains.(A–C) Representative conformations of docked CRD dimers. The figures illustrate the conformations showing the lowest intermolecular S-S distance after homophilic (A,B) and heterophilic (C) association obtained through the molecular docking of N-CRD (A) and C-CRD (B) domains. (D–F) Distributions of the S-S minimal distances retrieved from the docking of N-CRD with N-CRD (D), C-CRD with C-CRD (E) and N-CRD with C-CRD (F) domains. In (G) the cumulative distributions relative to the histograms shown in (D–F) are reported. (I) The graph shows the decay of dimer as a function of time quantified from the graphical post-processing of the western blot image shown in (H). The sequestration of the dimer in solution is shown for the N-CRD (green) and C-CRD (red) homopolymerisation, for the N-CRD and C-CRD heteropolymerisation (orange) and for the C-CRD (C9/18A) mutant homopolymerisation (black). Solid lines show the fitting of each dataset performed using a single exponential decay model (y = e−x/τ).
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f2: Propensity of intermolecular disulfide formation revealed by molecular docking of N-CRD and C-CRD domains.(A–C) Representative conformations of docked CRD dimers. The figures illustrate the conformations showing the lowest intermolecular S-S distance after homophilic (A,B) and heterophilic (C) association obtained through the molecular docking of N-CRD (A) and C-CRD (B) domains. (D–F) Distributions of the S-S minimal distances retrieved from the docking of N-CRD with N-CRD (D), C-CRD with C-CRD (E) and N-CRD with C-CRD (F) domains. In (G) the cumulative distributions relative to the histograms shown in (D–F) are reported. (I) The graph shows the decay of dimer as a function of time quantified from the graphical post-processing of the western blot image shown in (H). The sequestration of the dimer in solution is shown for the N-CRD (green) and C-CRD (red) homopolymerisation, for the N-CRD and C-CRD heteropolymerisation (orange) and for the C-CRD (C9/18A) mutant homopolymerisation (black). Solid lines show the fitting of each dataset performed using a single exponential decay model (y = e−x/τ).

Mentions: To elucidate the molecular basis for the association preferences of the two domains observed experimentally, docking of N-CRDs and C-CRDs into homo- and heterodimers was performed. We used a flexible docking algorithm, which allowed conformational adaptations of side chains including cysteines upon complex formation. We then assessed the ability of each monomer to form inter-domain disulfides by measuring the minimal S-S distance between the docked CRDs. We obtained a large set of putative relative poses for N-N, C-C, and N-C complexes (Fig. 2). Both, N-CRD and C-CRD, were able to dock and form homodimers, in which the minimal interdomain S-S distance is compatible (<0.4 nm) with intermolecular reshuffling of S-S bonds (Fig. 2A,B,D,E). However, the C-CRD showed a more pronounced tendency to associate, as reflected by lower S-S intermolecular distances (Fig. 2B,E). The heterodimer formation showed an intermediate profile with a distribution of S-S minimal distances that includes the values encountered in the N-CRD and C-CRD homophilic docking (Fig. 2C,F). These differences are particularly evident in the cumulative S-S distance distributions (Fig. 2G).


Minicollagen cysteine-rich domains encode distinct modes of polymerization to form stable nematocyst capsules.

Tursch A, Mercadante D, Tennigkeit J, Gräter F, Özbek S - Sci Rep (2016)

Propensity of intermolecular disulfide formation revealed by molecular docking of N-CRD and C-CRD domains.(A–C) Representative conformations of docked CRD dimers. The figures illustrate the conformations showing the lowest intermolecular S-S distance after homophilic (A,B) and heterophilic (C) association obtained through the molecular docking of N-CRD (A) and C-CRD (B) domains. (D–F) Distributions of the S-S minimal distances retrieved from the docking of N-CRD with N-CRD (D), C-CRD with C-CRD (E) and N-CRD with C-CRD (F) domains. In (G) the cumulative distributions relative to the histograms shown in (D–F) are reported. (I) The graph shows the decay of dimer as a function of time quantified from the graphical post-processing of the western blot image shown in (H). The sequestration of the dimer in solution is shown for the N-CRD (green) and C-CRD (red) homopolymerisation, for the N-CRD and C-CRD heteropolymerisation (orange) and for the C-CRD (C9/18A) mutant homopolymerisation (black). Solid lines show the fitting of each dataset performed using a single exponential decay model (y = e−x/τ).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4863159&req=5

f2: Propensity of intermolecular disulfide formation revealed by molecular docking of N-CRD and C-CRD domains.(A–C) Representative conformations of docked CRD dimers. The figures illustrate the conformations showing the lowest intermolecular S-S distance after homophilic (A,B) and heterophilic (C) association obtained through the molecular docking of N-CRD (A) and C-CRD (B) domains. (D–F) Distributions of the S-S minimal distances retrieved from the docking of N-CRD with N-CRD (D), C-CRD with C-CRD (E) and N-CRD with C-CRD (F) domains. In (G) the cumulative distributions relative to the histograms shown in (D–F) are reported. (I) The graph shows the decay of dimer as a function of time quantified from the graphical post-processing of the western blot image shown in (H). The sequestration of the dimer in solution is shown for the N-CRD (green) and C-CRD (red) homopolymerisation, for the N-CRD and C-CRD heteropolymerisation (orange) and for the C-CRD (C9/18A) mutant homopolymerisation (black). Solid lines show the fitting of each dataset performed using a single exponential decay model (y = e−x/τ).
Mentions: To elucidate the molecular basis for the association preferences of the two domains observed experimentally, docking of N-CRDs and C-CRDs into homo- and heterodimers was performed. We used a flexible docking algorithm, which allowed conformational adaptations of side chains including cysteines upon complex formation. We then assessed the ability of each monomer to form inter-domain disulfides by measuring the minimal S-S distance between the docked CRDs. We obtained a large set of putative relative poses for N-N, C-C, and N-C complexes (Fig. 2). Both, N-CRD and C-CRD, were able to dock and form homodimers, in which the minimal interdomain S-S distance is compatible (<0.4 nm) with intermolecular reshuffling of S-S bonds (Fig. 2A,B,D,E). However, the C-CRD showed a more pronounced tendency to associate, as reflected by lower S-S intermolecular distances (Fig. 2B,E). The heterodimer formation showed an intermediate profile with a distribution of S-S minimal distances that includes the values encountered in the N-CRD and C-CRD homophilic docking (Fig. 2C,F). These differences are particularly evident in the cumulative S-S distance distributions (Fig. 2G).

Bottom Line: Our combined experimental and computational analyses reveal the cysteines in the C-CRD fold to exhibit a higher structural propensity for disulfide bonding and a faster kinetics of polymerization.During nematocyst maturation, the highly reactive C-CRD is instrumental in efficient cross-linking of minicollagens to form pressure resistant capsules.The higher ratio of C-CRD folding types evidenced in the medusozoan lineage might have fostered the evolution of novel, predatory nematocyst types in cnidarians with a free-swimming medusa stage.

View Article: PubMed Central - PubMed

Affiliation: University of Heidelberg, Centre for Organismal Studies, Department of Molecular Evolution and Genomics, Im Neuenheimer Feld 329, 69120 Heidelberg, Germany.

ABSTRACT
The stinging capsules of cnidarians, nematocysts, function as harpoon-like organelles with unusual biomechanical properties. The nanosecond discharge of the nematocyst requires a dense protein network of the capsule structure withstanding an internal pressure of up to 150 bar. Main components of the capsule are short collagens, so-called minicollagens, that form extended polymers by disulfide reshuffling of their cysteine-rich domains (CRDs). Although CRDs have identical cysteine patterns, they exhibit different structures and disulfide connectivity at minicollagen N and C-termini. We show that the structurally divergent CRDs have different cross-linking potentials in vitro and in vivo. While the C-CRD can participate in several simultaneous intermolecular disulfides and functions as a cystine knot after minicollagen synthesis, the N-CRD is monovalent. Our combined experimental and computational analyses reveal the cysteines in the C-CRD fold to exhibit a higher structural propensity for disulfide bonding and a faster kinetics of polymerization. During nematocyst maturation, the highly reactive C-CRD is instrumental in efficient cross-linking of minicollagens to form pressure resistant capsules. The higher ratio of C-CRD folding types evidenced in the medusozoan lineage might have fostered the evolution of novel, predatory nematocyst types in cnidarians with a free-swimming medusa stage.

No MeSH data available.


Related in: MedlinePlus