Nautilus pompilius Hemocyanin: 9 Å Cryo-EM Structure and Molecular Model Reveal the Subunit Pathway and the Interfaces between the 70 Functional Units

Gatsogiannis, Christos; Moeller, Arne; Depoix, Frank; Meissner, Ulrich; Markl, Jürgen

doi:10.1016/j.jmb.2007.09.036

Cited by 43 publications

(93 citation statements)

References 59 publications

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“…As a consequence, the subunit dimer of NpH is an asymmetric homodimer. 5 All NpH FUs are arranged pairwise as morphological units, and we identified the following duplets: a ↔ b, d ↔ e, c ↔ f, and g ↔ g. We also detected that the pairing mode strikingly resembles the protomer-protomer association in the published crystal lattice of FU-g from Octopus dofleini hemocyanin (OdH). 6 In side-view orientation of the decamer, the wall appears as three tiers, with the FU repeat a-b-c-f in the two peripheral tiers and with d-e-e-d in the central tier.…”

Section: Introductionmentioning

confidence: 70%

“…2a-d; for comparison to NpH, see Figs. 2 and 3 in Gatsogiannis et al 5 ). This similarity encompasses the shape and orientation of the six wall FU types, and specific wall features (such as the anchor, and Gatsogiannis et al 5 ).…”

Section: Resultsmentioning

confidence: 94%

“…5 Our present report on keyhole limpet hemocyanin (KLH) 1 should be primarily considered alongside this NpH study. We demonstrated that this cephalopod hemocyanin is composed of 10 identical 340-kDa subunits arranged as anti-parallel pairs; due to an oblique arrangement of the five subunit dimers, the cylinder wall appears as a right-handed helix.…”

Section: Introductionmentioning

confidence: 99%

“…From these data, NpH can be considered as a molluscan hemocyanin prototype. 5 Gastropod hemocyanins such as KLH1 differ from hemocyanin molecules of the NpH prototype in four major aspects: (i) the native molecule is a didecamer, with two decamers assembled face-toface at their open ends; 1 (ii) the gastropod hemocyanin subunit is a 400-kDa polypeptide due to the presence of an additional FU type (FU-h); (iii) the asymmetric collar complex of the decameric halfmolecule is enlarged by an outer annulus of five "slabs" representing FU-h pairs; 7 and (iv) FU-h carries a C-terminal tail of ca 100 amino acids 8 that has recently been found to possess a cupredoxinlike fold important for h ↔ h anti-parallel pairing. 9 Our recent comparative cryoEM three-dimensional (3D) analysis at 11-Å resolution has shown that the wall of Nautilus hemocyanin is very similar to the wall of a gastropod hemocyanin [Haliotis tuberculata hemocyanin (HtH) 1], but the two collar complexes appeared to be substantially different; however, at 11-Å, a reliable molecular model of the didecamer could not be obtained.…”

Section: Introductionmentioning

confidence: 99%

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Keyhole Limpet Hemocyanin: 9-Å CryoEM Structure and Molecular Model of the KLH1 Didecamer Reveal the Interfaces and Intricate Topology of the 160 Functional Units

Gatsogiannis

Markl

2009

Journal of Molecular Biology

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115

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Section: Introductionmentioning

confidence: 70%

Section: Resultsmentioning

confidence: 94%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Keyhole Limpet Hemocyanin: 9-Å CryoEM Structure and Molecular Model of the KLH1 Didecamer Reveal the Interfaces and Intricate Topology of the 160 Functional Units

Gatsogiannis

Markl

2009

Journal of Molecular Biology

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115

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“…These cylinders either exist as decamers or aggregate further to form didecamers or multidecamers. The mollusc hemocyanin subunit has a molecular mass of 350-400 kDa and is a concatenation of seven or eight paralogous FUs of approximately 50 kDa each (FU-a to FU-h) on a single polypeptide chain (2,16,17). Each FU is typically made-up from two domains and bears one active site.…”

Section: Introductionmentioning

confidence: 99%

The refined structure of functional unit h of keyhole limpet hemocyanin (KLH1‐h) reveals disulfide bridges

et al. 2011

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Hemocyanins are multimeric oxygen‐transport proteins in the hemolymph of many arthropods and mollusks. The overall molecular architecture of arthropod and molluscan hemocyanin is very different, although they possess a similar binuclear type 3 copper center to bind oxygen in a side‐on conformation. Gastropod hemocyanin is a 35 nm cylindrical didecamer (2 × 10‐mer) based on a 400 kDa subunit. The latter is subdivided into eight paralogous “functional units” (FU‐a to FU‐h), each with an active site. FU‐a to FU‐f contribute to the cylinder wall, whereas FU‐g and FU‐h form the internal collar complex. Atomic structures of FU‐e and FU‐g, and a 9 Å cryoEM structure of the 8 MDa didecamer are available. Recently, the structure of keyhole limpet hemocyanin FU‐h (KLH1‐h) was presented as a Cα‐trace at 4 Å resolution. Unlike the other seven FU types, FU‐h contains an additional C‐terminal domain with a cupredoxin‐like fold. Because of the resolution limit of 4 Å, in some loops, the course of the protein backbone could not be established with high certainty yet. Here, we present a refined atomic structure of FU‐h (KLH1‐h) obtained from low‐resolution refinement, which unambiguously establishes the course of the polypeptide backbone and reveals the disulfide bridges as well as the orientation of bulky amino acids. © 2011 IUBMB IUBMB Life, 63(3): 183–187, 2011

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ESI‐MS and MALLS analysis of quaternary structure of molluscan hemocyanins

et al. 2012

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The understanding of the function of macromolecular complexes is mainly related to a precise knowledge of their structure. Recently, the development of suitable mass spectrometric techniques (electrospray ionization (ESI) and matrix-assisted laser desorption/ionization (MALDI)) and multi-angle laser light scattering has enabled mass determination of native complexes and of their subunits. By these techniques, the structure and association/dissociation behavior of huge molecules of molluscan Octopus vulgaris, Sepia officinalis and Rapana venosa have been characterized. Molecular masses of the native and dissociated molecule of cephalopodan Hcs O. vulgaris (3545 and 359.3 kDa, respectively) and S. officinalis (4134 and 443.8 kDa, respectively) revealed that only one type subunit organizes their molecules, while the presence of two isoforms with different masses (422.8 and 400.0 kDa) has been determined for gastropodan R. venosa Hc, aggregated into didecamers. The difference of their structural subunits was also established after limited proteolysis with TPCK-trypsin. Eight functional units (FUs) with masses of ~ 50 kDa were isolated from both subunits of RvH and isoform of Sepia officinalis, while seven FUs were purified from OvH. Further characterization of proteins by ESI-mass spectrometry (MS) and MALDI-MS, methods gave insights into post-translational modifications such as glycosylation. Glycosylation of O. vulgaris and S. officinalis Hcs was suggested based on the differences (11.6 and 40.0 kDa, respectively) between the masses measured by ESI-MS and those calculated by their gene sequences.

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Nautilus pompilius Hemocyanin: 9 Å Cryo-EM Structure and Molecular Model Reveal the Subunit Pathway and the Interfaces between the 70 Functional Units

Cited by 43 publications

References 59 publications

Keyhole Limpet Hemocyanin: 9-Å CryoEM Structure and Molecular Model of the KLH1 Didecamer Reveal the Interfaces and Intricate Topology of the 160 Functional Units

Keyhole Limpet Hemocyanin: 9-Å CryoEM Structure and Molecular Model of the KLH1 Didecamer Reveal the Interfaces and Intricate Topology of the 160 Functional Units

The refined structure of functional unit h of keyhole limpet hemocyanin (KLH1‐h) reveals disulfide bridges

ESI‐MS and MALLS analysis of quaternary structure of molluscan hemocyanins

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