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Discovery of a distinct superfamily of Kunitz-type toxin (KTT) from tarantulas.

Yuan CH, He QY, Peng K, Diao JB, Jiang LP, Tang X, Liang SP - PLoS ONE (2008)

Bottom Line: Kuntiz-type toxins (KTTs) have been found in the venom of animals such as snake, cone snail and sea anemone.The results also revealed a series of key events in the history of spider KTT evolution, including the formation of a novel KTT family (named sub-Kuntiz-type toxins) derived from the ancestral native KTTs with the loss of the second disulfide bridge accompanied by several dramatic sequence modifications.These finding illustrate that the two activity sites of Kunitz-type toxins are functionally and evolutionally independent and provide new insights into effects of Darwinian selection pressures on KTT evolution, and mechanisms by which new functions can be grafted onto old protein scaffolds.

View Article: PubMed Central - PubMed

Affiliation: The Key Laboratory for Protein Chemistry and Developmental Biology of Ministry of Education, College of Life Sciences, Hunan Normal University, Changsha, PR China.

ABSTRACT

Background: Kuntiz-type toxins (KTTs) have been found in the venom of animals such as snake, cone snail and sea anemone. The main ancestral function of Kunitz-type proteins was the inhibition of a diverse array of serine proteases, while toxic activities (such as ion-channel blocking) were developed under a variety of Darwinian selection pressures. How new functions were grafted onto an old protein scaffold and what effect Darwinian selection pressures had on KTT evolution remains a puzzle.

Principal findings: Here we report the presence of a new superfamily of ktts in spiders (TARANTULAS: Ornithoctonus huwena and Ornithoctonus hainana), which share low sequence similarity to known KTTs and is clustered in a distinct clade in the phylogenetic tree of KTT evolution. The representative molecule of spider KTTs, HWTX-XI, purified from the venom of O. huwena, is a bi-functional protein which is a very potent trypsin inhibitor (about 30-fold more strong than BPTI) as well as a weak Kv1.1 potassium channel blocker. Structural analysis of HWTX-XI in 3-D by NMR together with comparative function analysis of 18 expressed mutants of this toxin revealed two separate sites, corresponding to these two activities, located on the two ends of the cone-shape molecule of HWTX-XI. Comparison of non-synonymous/synonymous mutation ratios (omega) for each site in spider and snake KTTs, as well as PBTI like body Kunitz proteins revealed high Darwinian selection pressure on the binding sites for Kv channels and serine proteases in snake, while only on the proteases in spider and none detected in body proteins, suggesting different rates and patterns of evolution among them. The results also revealed a series of key events in the history of spider KTT evolution, including the formation of a novel KTT family (named sub-Kuntiz-type toxins) derived from the ancestral native KTTs with the loss of the second disulfide bridge accompanied by several dramatic sequence modifications.

Conclusions/significance: These finding illustrate that the two activity sites of Kunitz-type toxins are functionally and evolutionally independent and provide new insights into effects of Darwinian selection pressures on KTT evolution, and mechanisms by which new functions can be grafted onto old protein scaffolds.

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Phylogenetic tree of the KTTs and nonsynonymous/synonymous mutation ratios (ω) comparison.(A), A minimum evolution (ME) tree of mature fragments of KTTs and some Kunitz type body proteins in various taxon groups. The branches are coloured as follows: red, spider KTTs; cyan, sea anemone; green, BPTI homologies from mammals; dark yellow, snake KTTs; blue, conkunitzin from cone snail; black, others. (B), Column charts of the posterior mean nonsynonymous/synonymous mutation ratios (ω) for each site in spider, snake KTTs as well as body Kunitz proteins. In these charts, the deep colour column is based on Model 3 (discrete) and the light one is based on Model 8 (beta&ω>1). Region A and B are marked by yellow and cyan backgrounds, and the secondary structure of HWTX-XI is shown on the bottom. (C), Surface models of representative spider KTTs (HWTX-XI), snake KTTs (DTXK) and Body proteins (BPTI). The residues are colored based on the value of ω in Model 8.
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pone-0003414-g009: Phylogenetic tree of the KTTs and nonsynonymous/synonymous mutation ratios (ω) comparison.(A), A minimum evolution (ME) tree of mature fragments of KTTs and some Kunitz type body proteins in various taxon groups. The branches are coloured as follows: red, spider KTTs; cyan, sea anemone; green, BPTI homologies from mammals; dark yellow, snake KTTs; blue, conkunitzin from cone snail; black, others. (B), Column charts of the posterior mean nonsynonymous/synonymous mutation ratios (ω) for each site in spider, snake KTTs as well as body Kunitz proteins. In these charts, the deep colour column is based on Model 3 (discrete) and the light one is based on Model 8 (beta&ω>1). Region A and B are marked by yellow and cyan backgrounds, and the secondary structure of HWTX-XI is shown on the bottom. (C), Surface models of representative spider KTTs (HWTX-XI), snake KTTs (DTXK) and Body proteins (BPTI). The residues are colored based on the value of ω in Model 8.

Mentions: Phylogenetic analyses based on comparison of mature spider KTTs with those inother taxonomic groups, including sea anemone, snake and some body PBTI like proteins [16], [17], [24] has demonstrated that all known Kunitz proteins can be clustered into three clades at the top level and that the spider KTTs are restricted to a clade which appears between the snake and primitive Kunitz type protein clades (Fig. 9A and Text S2). The branch lengths of spider toxins are much longer than that of snake KTTs, suggesting an earlier recruitment event for spider toxins than for snake toxins.


Discovery of a distinct superfamily of Kunitz-type toxin (KTT) from tarantulas.

Yuan CH, He QY, Peng K, Diao JB, Jiang LP, Tang X, Liang SP - PLoS ONE (2008)

Phylogenetic tree of the KTTs and nonsynonymous/synonymous mutation ratios (ω) comparison.(A), A minimum evolution (ME) tree of mature fragments of KTTs and some Kunitz type body proteins in various taxon groups. The branches are coloured as follows: red, spider KTTs; cyan, sea anemone; green, BPTI homologies from mammals; dark yellow, snake KTTs; blue, conkunitzin from cone snail; black, others. (B), Column charts of the posterior mean nonsynonymous/synonymous mutation ratios (ω) for each site in spider, snake KTTs as well as body Kunitz proteins. In these charts, the deep colour column is based on Model 3 (discrete) and the light one is based on Model 8 (beta&ω>1). Region A and B are marked by yellow and cyan backgrounds, and the secondary structure of HWTX-XI is shown on the bottom. (C), Surface models of representative spider KTTs (HWTX-XI), snake KTTs (DTXK) and Body proteins (BPTI). The residues are colored based on the value of ω in Model 8.
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Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2561067&req=5

pone-0003414-g009: Phylogenetic tree of the KTTs and nonsynonymous/synonymous mutation ratios (ω) comparison.(A), A minimum evolution (ME) tree of mature fragments of KTTs and some Kunitz type body proteins in various taxon groups. The branches are coloured as follows: red, spider KTTs; cyan, sea anemone; green, BPTI homologies from mammals; dark yellow, snake KTTs; blue, conkunitzin from cone snail; black, others. (B), Column charts of the posterior mean nonsynonymous/synonymous mutation ratios (ω) for each site in spider, snake KTTs as well as body Kunitz proteins. In these charts, the deep colour column is based on Model 3 (discrete) and the light one is based on Model 8 (beta&ω>1). Region A and B are marked by yellow and cyan backgrounds, and the secondary structure of HWTX-XI is shown on the bottom. (C), Surface models of representative spider KTTs (HWTX-XI), snake KTTs (DTXK) and Body proteins (BPTI). The residues are colored based on the value of ω in Model 8.
Mentions: Phylogenetic analyses based on comparison of mature spider KTTs with those inother taxonomic groups, including sea anemone, snake and some body PBTI like proteins [16], [17], [24] has demonstrated that all known Kunitz proteins can be clustered into three clades at the top level and that the spider KTTs are restricted to a clade which appears between the snake and primitive Kunitz type protein clades (Fig. 9A and Text S2). The branch lengths of spider toxins are much longer than that of snake KTTs, suggesting an earlier recruitment event for spider toxins than for snake toxins.

Bottom Line: Kuntiz-type toxins (KTTs) have been found in the venom of animals such as snake, cone snail and sea anemone.The results also revealed a series of key events in the history of spider KTT evolution, including the formation of a novel KTT family (named sub-Kuntiz-type toxins) derived from the ancestral native KTTs with the loss of the second disulfide bridge accompanied by several dramatic sequence modifications.These finding illustrate that the two activity sites of Kunitz-type toxins are functionally and evolutionally independent and provide new insights into effects of Darwinian selection pressures on KTT evolution, and mechanisms by which new functions can be grafted onto old protein scaffolds.

View Article: PubMed Central - PubMed

Affiliation: The Key Laboratory for Protein Chemistry and Developmental Biology of Ministry of Education, College of Life Sciences, Hunan Normal University, Changsha, PR China.

ABSTRACT

Background: Kuntiz-type toxins (KTTs) have been found in the venom of animals such as snake, cone snail and sea anemone. The main ancestral function of Kunitz-type proteins was the inhibition of a diverse array of serine proteases, while toxic activities (such as ion-channel blocking) were developed under a variety of Darwinian selection pressures. How new functions were grafted onto an old protein scaffold and what effect Darwinian selection pressures had on KTT evolution remains a puzzle.

Principal findings: Here we report the presence of a new superfamily of ktts in spiders (TARANTULAS: Ornithoctonus huwena and Ornithoctonus hainana), which share low sequence similarity to known KTTs and is clustered in a distinct clade in the phylogenetic tree of KTT evolution. The representative molecule of spider KTTs, HWTX-XI, purified from the venom of O. huwena, is a bi-functional protein which is a very potent trypsin inhibitor (about 30-fold more strong than BPTI) as well as a weak Kv1.1 potassium channel blocker. Structural analysis of HWTX-XI in 3-D by NMR together with comparative function analysis of 18 expressed mutants of this toxin revealed two separate sites, corresponding to these two activities, located on the two ends of the cone-shape molecule of HWTX-XI. Comparison of non-synonymous/synonymous mutation ratios (omega) for each site in spider and snake KTTs, as well as PBTI like body Kunitz proteins revealed high Darwinian selection pressure on the binding sites for Kv channels and serine proteases in snake, while only on the proteases in spider and none detected in body proteins, suggesting different rates and patterns of evolution among them. The results also revealed a series of key events in the history of spider KTT evolution, including the formation of a novel KTT family (named sub-Kuntiz-type toxins) derived from the ancestral native KTTs with the loss of the second disulfide bridge accompanied by several dramatic sequence modifications.

Conclusions/significance: These finding illustrate that the two activity sites of Kunitz-type toxins are functionally and evolutionally independent and provide new insights into effects of Darwinian selection pressures on KTT evolution, and mechanisms by which new functions can be grafted onto old protein scaffolds.

Show MeSH
Related in: MedlinePlus