Solution structures of 2 × 6-meric and 4 × 6-meric hemocyanins of crustaceans Carcinus aestuarii, Squilla mantis and Upogebia pusilla

Mičetić, Ivan; Losasso, Carmen; Muro, Paolo Di; Tognon, Giuseppe; Benedetti, Piero; Beltramini, Mariano

doi:10.1016/j.jsb.2010.03.012

Cited by 7 publications

(7 citation statements)

References 41 publications

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“…The five, previously described β 3 oligomers were used as starting points for the building of αβγ‐heterotrimers by substitution of the subunits with homology‐modelled α and γ structures. The use of modelled subunits is justified because the structural error in amino acid placement during homology modelling is smaller than the error obtained by SAXS‐based rigid body modelling [28]. Ten heterotrimers were thus created (five αβγ and five γβα trimers).…”

Section: Resultsmentioning

confidence: 99%

Structural analysis of trimeric phospholipase A₂ neurotoxin from the Australian taipan snake venom

et al. 2012

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Snake pre-synaptic neurotoxins endowed with phospholipase A 2 activity are potent inducers of paralysis through the specific disruption of the neuromuscular junction pre-synaptic membrane and represent a valuable tool for investigating neuronal degeneration and recovery. They have different structural complexity and a wide range of lethal potency and enzymatic activity, although they share a similar mechanism of action. Although no correlation has been reported between neurotoxicity and enzymatic activity, toxicity increases with structural complexity and phospholipase A 2 oligomers show 10-fold lower LD 50 values compared to their monomeric counterparts. To date, no structural study has been performed on multimeric SPANs with the aim of shedding light on the correlation between structural complexity and neurotoxicity. In the present study, we investigated the structure of taipoxin, a trimeric phospholipase A 2 neurotoxin, as well as that of its subunits, by X-ray crystallography and small angle X-ray scattering analysis. We present the high-resolution structure of two isoforms of the taipoxin b subunit, which show no neurotoxic activity but enhance the activity of the other subunits in the complex. One isoform shows no structural change that could justify the lack of activity. The other displays three point mutations in critical positions for the catalytic activity. Moreover, we designed a model for the quaternary structure of taipoxin under physiological conditions, in which the three subunits are organized into a flat holotoxin with the substrate binding sockets exposed on the same side of the complex, which suggests a role for this interface in the toxin-membrane interaction. DatabaseThe coordinates and structure factors have been deposited in the RCSB Protein Data Bank (http://www.rcsb.org) under accession numbers 3VBZ and 3VC0, corresponding to b isoforms 1 and 2 respectively Structure digital abstract l taipoxin beta isoform 2 and taipoxin beta isoform 2 bind by x-ray crystallography (View interaction)Abbreviations CGN, cerebellar granule neurone; PDB, Protein Data Bank; PLA 2, phospholipase A 2; SAXS, small angle X-ray scattering; SPAN, snake phospholipase A 2 neurotoxin.

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Section: Resultsmentioning

confidence: 99%

Structural analysis of trimeric phospholipase A₂ neurotoxin from the Australian taipan snake venom

et al. 2012

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“…Hemocyanins are copper-containing respiratory proteins, dissolved in concentrations up to 120 mg/ml in the hemolymph of molluscs and arthropods, including Crustacea, Chelicerata, Myriapoda, and some species of insects (Hagner-Holler et al 2004;Sánchez et al 1998;van Holde and Miller 1995;Chen et al 2007;Mičetic et al 2010). These molecules are composed of a number of subunits that assemble in an extremely large macro-molecular entity.…”

Section: Introductionmentioning

confidence: 99%

Structure and Characterization of Eriphia verrucosa Hemocyanin

et al. 2015

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Arthropod hemocyanins (Hcs) are a family of large extracellular oxygen-transporting proteins with high molecular mass and hexameric or multi-hexameric molecular assembly. This study reports for the first time the isolation and characterization of the structure of an arthropod hemocyanin from crab Eriphia verrucosa (EvH) living in the Black Sea. Its oligomeric quaternary structure is based on different arrangements of a basic 6 × 75 kDa hexameric unit, and four of them (EvH1, EvH2, EvH3, and EvH4) were identified using ion-exchange chromatography. Subunit 3 (EvH3) shows high similarity scores (75.0, 87.5, 91.7, and 75.0 %, respectively) by comparison of the N-terminal sequence of subunit 1 from Cancer pagurus of the North Sea (Cp1), subunits 3 and 6 of Cancer magister (Cm3 and Cm6), and subunit 2 of Carcinus aestuarii (CaSS2), respectively. Moreover, a partial cDNA sequence (1309 bp) of E. verrucosa hemocyanin encoding a protein of 435 amino acids was isolated. The deduced amino acid sequence shows a high degree of similarity with subunits 3, 4, 5, and 6 of C. magister (81-84 %). Most of the hemocyanins are glycosylated, and three putative O-linkage sites were identified in the partial amino acid sequence of EvH at positions 444-446, 478-480, and 547-549, respectively. The higher stability of native Hc in comparison to its subunit EvH4 as determined by circular dichroism (CD) could be explained with the formation of a stabilizing quaternary structure.

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“…Proteome maps of the hepatopancreas of crustacean C. pagurus were performed by 2D-PAGE, and vitellogenin (yolk protein), several subunits of hemocyanin, cryptocyanin, ferritin, and calreticulin were identified (Ward et al 2010). Hemocyanins are coppercontaining respiratory proteins, dissolved in concentrations up to 120 mg/ml in the hemolymph of molluscs and arthropods, including Crustacea, Chelicerata, Myriapoda, and some species of insects (Hagner-Holler et al 2004;Sánchez et al 1998;van Holde and Miller 1995;Chen et al 2007;Mičetic et al 2010). These molecules are composed of a number of subunits that assemble in an extremely large macro-molecular entity.…”

Section: Introductionmentioning

confidence: 99%

RETRACTED ARTICLE: Structure and Characterization of Cancer pagurus Hemocyanin

et al. 2014

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Solution structures of 2 × 6-meric and 4 × 6-meric hemocyanins of crustaceans Carcinus aestuarii, Squilla mantis and Upogebia pusilla

Cited by 7 publications

References 41 publications

Structural analysis of trimeric phospholipase A₂ neurotoxin from the Australian taipan snake venom

Structural analysis of trimeric phospholipase A₂ neurotoxin from the Australian taipan snake venom

Structure and Characterization of Eriphia verrucosa Hemocyanin

RETRACTED ARTICLE: Structure and Characterization of Cancer pagurus Hemocyanin

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Solution structures of 2 × 6-meric and 4 × 6-meric hemocyanins of crustaceans Carcinus aestuarii, Squilla mantis and Upogebia pusilla

Cited by 7 publications

References 41 publications

Structural analysis of trimeric phospholipase A2 neurotoxin from the Australian taipan snake venom

Structural analysis of trimeric phospholipase A2 neurotoxin from the Australian taipan snake venom

Structure and Characterization of Eriphia verrucosa Hemocyanin

RETRACTED ARTICLE: Structure and Characterization of Cancer pagurus Hemocyanin

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Product

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Structural analysis of trimeric phospholipase A₂ neurotoxin from the Australian taipan snake venom

Structural analysis of trimeric phospholipase A₂ neurotoxin from the Australian taipan snake venom