A new multigene superfamily of Kunitz-type protease inhibitors from sea anemone Heteractis crispa

Isaeva, Marina; Chausova, Victoria; Zelepuga, Elena; Guzev, Konstantin V.; Tabakmakher, Valentin М.; Monastyrnaya, Margarita; Kozlovskaya, É. P.

doi:10.1016/j.peptides.2011.09.022

Cited by 41 publications

(55 citation statements)

References 54 publications

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“…Here for, nested PCR with gene specific primers created on the basis of nucleotide sequences of Kunitz-type peptide genes was used [11] (Table S1). Analysis of the deduced amino acid sequences revealed that all peptides have a N-terminal Arg1 and Lys14 at the P1 position.…”

Section: Resultsmentioning

confidence: 99%

“…The peptides of this family have an ancient Kunitz fold and some of them are characterized by a unique and intriguing feature of dual functionality since they inhibit both proteases and ion channels [7,8,9,10]. This phenomenon is probably caused by the selection of peptide structures during divergent evolution together with specialization, diversification and sub- and neofunctionalization [11]. Considering an increasing diversity in sea anemone preys and predators, more than one function in the same peptide scaffold is advantageous for the survival of these organisms on Earth for nearly 500 million years [12].…”

Section: Introductionmentioning

confidence: 99%

“…It was found that H. crispa Kunitz-type protease inhibitors are coded by the multigene superfamily and form a combinatorial library including HCGS, HCRG, HCGN, and HCGG peptide subfamilies [11]. The HCGS peptide subfamily and their evolutionary relationships were described in details previously [11,25].…”

Section: Introductionmentioning

confidence: 99%

“…The HCGS peptide subfamily and their evolutionary relationships were described in details previously [11,25]. Two sea anemone trypsin inhibitors, RmInI and RmInII, possess antihistamine activity in vivo [15].…”

Section: Introductionmentioning

confidence: 99%

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Kunitz-Type Peptide HCRG21 from the Sea Anemone Heteractis crispa Is a Full Antagonist of the TRPV1 Receptor

et al. 2016

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Sea anemone venoms comprise multifarious peptides modulating biological targets such as ion channels or receptors. The sequence of a new Kunitz-type peptide, HCRG21, belonging to the Heteractis crispa RG (HCRG) peptide subfamily was deduced on the basis of the gene sequence obtained from the Heteractis crispa cDNA. HCRG21 shares high structural homology with Kunitz-type peptides APHC1–APHC3 from H. crispa, and clusters with the peptides from so named “analgesic cluster” of the HCGS peptide subfamily but forms a separate branch on the NJ-phylogenetic tree. Three unique point substitutions at the N-terminus of the molecule, Arg1, Gly2, and Ser5, distinguish HCRG21 from other peptides of this cluster. The trypsin inhibitory activity of recombinant HCRG21 (rHCRG21) was comparable with the activity of peptides from the same cluster. Inhibition constants for trypsin and α-chymotrypsin were 1.0 × 10−7 and 7.0 × 10−7 M, respectively. Electrophysiological experiments revealed that rHCRG21 inhibits 95% of the capsaicin-induced current through transient receptor potential family member vanilloid 1 (TRPV1) and has a half-maximal inhibitory concentration of 6.9 ± 0.4 μM. Moreover, rHCRG21 is the first full peptide TRPV1 inhibitor, although displaying lower affinity for its receptor in comparison with other known ligands. Macromolecular docking and full atom Molecular Dynamics (MD) simulations of the rHCRG21–TRPV1 complex allow hypothesizing the existence of two feasible, intra- and extracellular, molecular mechanisms of blocking. These data provide valuable insights in the structural and functional relationships and pharmacological potential of bifunctional Kunitz-type peptides.

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Section: Resultsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Kunitz-Type Peptide HCRG21 from the Sea Anemone Heteractis crispa Is a Full Antagonist of the TRPV1 Receptor

et al. 2016

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“…Notably, they have also been identified previously in cnidarian taxa. In the venom arsenal of sea anemones they exhibit a plesiotypic activity of trypsin-inhibition, but possess additional apotypic activities including potassium channel blockage [31,76,78,277,278]. BLAST searches of the Kunitz peptide detected in this study A mono domain kazal serine protease inhibitor with homology to coral Melithaea caledonica (UniProt accession number P82968) was discovered in the oral arm and tentacle venoms (Figure 4-12).…”

Section: Serine Protease Inhibitorsmentioning

confidence: 81%

Evolution and diversification of the cnidarian venom system

Jouiaei¹

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The phylum Cnidaria (corals, sea pens, sea anemones, jellyfish and hydroids) is the oldest venomous animal lineage (~750 million years old), making it an ideal phylum to understand the origin and diversification of venom. Cnidarians are characterised by specialised cellular structures called cnidae, which they utilise to inject mixtures of bioactive compounds or venom for predation and defence. In recent years cnidarian venoms have begun to be investigated as a potential source of novel bioactive therapeutics. However, in comparison with several venomous lineages that are relatively younger, such as snakes, cone snails and spiders, the diversification and molecular evolutionary regimes of toxins encoded by these fascinating and ancient animals remain poorly understood.The first experimental part of this thesis (Chapter 2) is comprised of the phylogenetic and molecular evolutionary histories of pharmacologically characterised cnidarian toxin families. These include peptide neurotoxins (voltage-gated Na + and K + channel-targeting toxins: NaTxs and KTxs, respectively), pore-forming toxins (actinoporins, aerolysin-related toxins, and jellyfish toxins) and small cysteine-rich peptides (SCRiPs). Our analyses show that most cnidarian toxins remain conserved under the strong influence of negative selection. A deeper analysis of the molecular evolutionary histories of neurotoxins demonstrates that type III KTxs have evolved from NaTxs under the regime of positive selection and likely represent a unique evolutionary innovation of the Actinioidea lineage. We also identify a few functionally important sites in other classes of neurotoxins and pore-forming toxins that experienced episodic adaptation. In addition, we describe the first family of neurotoxic peptides (SCRiPs) in reef-building corals, Acropora millepora, that are likely involved in the envenoming function.The third chapter describes the development of a novel venom extraction technique which is rapid, repeatable and cost effective. This technique involves using ethanol to both induce cnidae discharge and to recover venom contents in one step. Our model species is the notorious Australian box jellyfish (Chironex fleckeri) which has a notable impact on public health. By using the complementary approaches of transcriptomics, proteomics, mass spectrometry, histology, and scanning electron microscopy, this study compares the proteome profile of the new ethanol recovery based method to a previously reported protocol, based upon density purified intact cnidae and pressure induced disruption. This work not only results in the expansion of identified Australian box jellyfish venom components but also illustrates that ethanol extraction method could greatly augment future cnidarian venom proteomics research efforts.The forth chapter is constituted by previously described venom extraction technique, ethanolinduced discharge of cnidae, to rapidly and efficiently characterise venom components of the Jelly Blubber or Blue Blubber Jellyfish, Catostylus mosaicus. By us...

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Expression and characterization of Flavikunin: A Kunitz‐type serine protease inhibitor identified in the venom gland cDNA library of Bungarus flaviceps

Kaur

Devi

Saikia

et al. 2018

J Biochem & Molecular Tox

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Trancriptomic analysis of the venom gland cDNA library of Bungarus flaviceps revealed Kunitz‐type serine protease inhibitor as one of the major venom protein families with three groups A, B, C. One of the group B isoforms named Flavikunin, which lacked an extra cysteine residue involved in disulfide bond formation in β‐bungarotoxin, was synthesized, cloned, and overexpressed in Escherichia coli. To decipher the structure‐function relationship, the P1 residue of Flavikunin, histidine, was mutated to alanine and arginine. Purified wild‐type and mutant Flavikunins were screened against serine proteases‐thrombin, factor Xa, trypsin, chymotrypsin, plasmin, and elastase. The wild‐type and mutant Flavikunin (H∆R) inhibited plasmin with an IC 50 of 0.48 and 0.35 µM, respectively. The in‐silico study showed that P1 residue of wild‐type and mutant (H∆R) Flavikunin interacted with S1′ and S1 site of plasmin, respectively. Thus, histidine at the P1 position was found to be involved in plasmin inhibition with mild anticoagulant activity.

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A new multigene superfamily of Kunitz-type protease inhibitors from sea anemone Heteractis crispa

Cited by 41 publications

References 54 publications

Kunitz-Type Peptide HCRG21 from the Sea Anemone Heteractis crispa Is a Full Antagonist of the TRPV1 Receptor

Kunitz-Type Peptide HCRG21 from the Sea Anemone Heteractis crispa Is a Full Antagonist of the TRPV1 Receptor

Evolution and diversification of the cnidarian venom system

Expression and characterization of Flavikunin: A Kunitz‐type serine protease inhibitor identified in the venom gland cDNA library of Bungarus flaviceps

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