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Tryptogalinin is a tick Kunitz serine protease inhibitor with a unique intrinsic disorder.

Valdés JJ, Schwarz A, Cabeza de Vaca I, Calvo E, Pedra JH, Guallar V, Kotsyfakis M - PLoS ONE (2013)

Bottom Line: Using homology-based modeling (and other protein prediction programs) we were able to model and explain the multifaceted function of tryptogalinin.The N-terminus of the modeled tryptogalinin is detached from the rest of the peptide and exhibits intrinsic disorder allowing an increased flexibility for its high affinity with its inhibiting partners (i.e., serine proteases).By incorporating experimental and computational methods our data not only describes the function of a Kunitz peptide from Ixodes scapularis, but also allows us to hypothesize about the molecular basis of this function at the atomic level.

View Article: PubMed Central - PubMed

Affiliation: Institute of Parasitology, Biology Centre of the Academy of Sciences of the Czech Republic, České Budějovice, Czech Republic. valdjj@gmail.com

ABSTRACT

Background: A salivary proteome-transcriptome project on the hard tick Ixodes scapularis revealed that Kunitz peptides are the most abundant salivary proteins. Ticks use Kunitz peptides (among other salivary proteins) to combat host defense mechanisms and to obtain a blood meal. Most of these Kunitz peptides, however, remain functionally uncharacterized, thus limiting our knowledge about their biochemical interactions.

Results: We discovered an unusual cysteine motif in a Kunitz peptide. This peptide inhibits several serine proteases with high affinity and was named tryptogalinin due to its high affinity for β-tryptase. Compared with other functionally described peptides from the Acari subclass, we showed that tryptogalinin is phylogenetically related to a Kunitz peptide from Rhipicephalus appendiculatus, also reported to have a high affinity for β-tryptase. Using homology-based modeling (and other protein prediction programs) we were able to model and explain the multifaceted function of tryptogalinin. The N-terminus of the modeled tryptogalinin is detached from the rest of the peptide and exhibits intrinsic disorder allowing an increased flexibility for its high affinity with its inhibiting partners (i.e., serine proteases).

Conclusions: By incorporating experimental and computational methods our data not only describes the function of a Kunitz peptide from Ixodes scapularis, but also allows us to hypothesize about the molecular basis of this function at the atomic level.

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A–B. Phylogenetic reconstruction and sequence analysis of tryptogalinin and functionally described Kunitz from the literature.A phylogram (A) of tryptogalinin and other functionally described Kunitz peptides from hematophagous arthropods, nematodes and platyhelminthes was constructed using maximum likelihod (ML) methods. The first four letters for each label displays the taxa name followed by the GenBank accession number and the functional nomenclature from the literature. The presented ML tree was rooted with Hg1, a trypsin and potassium channel inhibitor of the Mexican scorpion Hadrurus gertschi (GenBank: P0C8W3). Bootstrap values (≥50%) are indicated in the phylogram and the scale bar (mean amino acid substitution/site) is presented at the bottom left corner. A sequence alignment (B) of tryptogalinin and TdPI of Rhipicephalus appendiculatus from clade IV. Identical (black) and similar sequence regions (grey), the positions of the conserved Cys residues (roman numerals and brackets), and the Cys-Lys-Ala sequence (box) that form the Kunitz head (P1) interacting with the active site of serine proteases are marked. The signal peptide for tryptogalinin is highlighted in yellow.
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pone-0062562-g004: A–B. Phylogenetic reconstruction and sequence analysis of tryptogalinin and functionally described Kunitz from the literature.A phylogram (A) of tryptogalinin and other functionally described Kunitz peptides from hematophagous arthropods, nematodes and platyhelminthes was constructed using maximum likelihod (ML) methods. The first four letters for each label displays the taxa name followed by the GenBank accession number and the functional nomenclature from the literature. The presented ML tree was rooted with Hg1, a trypsin and potassium channel inhibitor of the Mexican scorpion Hadrurus gertschi (GenBank: P0C8W3). Bootstrap values (≥50%) are indicated in the phylogram and the scale bar (mean amino acid substitution/site) is presented at the bottom left corner. A sequence alignment (B) of tryptogalinin and TdPI of Rhipicephalus appendiculatus from clade IV. Identical (black) and similar sequence regions (grey), the positions of the conserved Cys residues (roman numerals and brackets), and the Cys-Lys-Ala sequence (box) that form the Kunitz head (P1) interacting with the active site of serine proteases are marked. The signal peptide for tryptogalinin is highlighted in yellow.

Mentions: Since we showed that tryptogalanin inhibits several serine proteases, we were interested in the relationship of this protease inhibitor to other functionally described Kunitz peptides from hematophagous arthropods, nematodes and platyhelminthes. Our analysis showed that the overall phylogenetic relationship among these Kunitz peptides was not resolved using maximum likelihood (ML) methods, apparently due to their amino acid sequence diversity. Only a few internal clades in the ML tree showed bootstrap support values higher than 50%. The phylogram in Figure 4A reveals five well-supported clades of Kunitz protease inhibitors in soft and hard ticks, scorpions and horse flies.


Tryptogalinin is a tick Kunitz serine protease inhibitor with a unique intrinsic disorder.

Valdés JJ, Schwarz A, Cabeza de Vaca I, Calvo E, Pedra JH, Guallar V, Kotsyfakis M - PLoS ONE (2013)

A–B. Phylogenetic reconstruction and sequence analysis of tryptogalinin and functionally described Kunitz from the literature.A phylogram (A) of tryptogalinin and other functionally described Kunitz peptides from hematophagous arthropods, nematodes and platyhelminthes was constructed using maximum likelihod (ML) methods. The first four letters for each label displays the taxa name followed by the GenBank accession number and the functional nomenclature from the literature. The presented ML tree was rooted with Hg1, a trypsin and potassium channel inhibitor of the Mexican scorpion Hadrurus gertschi (GenBank: P0C8W3). Bootstrap values (≥50%) are indicated in the phylogram and the scale bar (mean amino acid substitution/site) is presented at the bottom left corner. A sequence alignment (B) of tryptogalinin and TdPI of Rhipicephalus appendiculatus from clade IV. Identical (black) and similar sequence regions (grey), the positions of the conserved Cys residues (roman numerals and brackets), and the Cys-Lys-Ala sequence (box) that form the Kunitz head (P1) interacting with the active site of serine proteases are marked. The signal peptide for tryptogalinin is highlighted in yellow.
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3643938&req=5

pone-0062562-g004: A–B. Phylogenetic reconstruction and sequence analysis of tryptogalinin and functionally described Kunitz from the literature.A phylogram (A) of tryptogalinin and other functionally described Kunitz peptides from hematophagous arthropods, nematodes and platyhelminthes was constructed using maximum likelihod (ML) methods. The first four letters for each label displays the taxa name followed by the GenBank accession number and the functional nomenclature from the literature. The presented ML tree was rooted with Hg1, a trypsin and potassium channel inhibitor of the Mexican scorpion Hadrurus gertschi (GenBank: P0C8W3). Bootstrap values (≥50%) are indicated in the phylogram and the scale bar (mean amino acid substitution/site) is presented at the bottom left corner. A sequence alignment (B) of tryptogalinin and TdPI of Rhipicephalus appendiculatus from clade IV. Identical (black) and similar sequence regions (grey), the positions of the conserved Cys residues (roman numerals and brackets), and the Cys-Lys-Ala sequence (box) that form the Kunitz head (P1) interacting with the active site of serine proteases are marked. The signal peptide for tryptogalinin is highlighted in yellow.
Mentions: Since we showed that tryptogalanin inhibits several serine proteases, we were interested in the relationship of this protease inhibitor to other functionally described Kunitz peptides from hematophagous arthropods, nematodes and platyhelminthes. Our analysis showed that the overall phylogenetic relationship among these Kunitz peptides was not resolved using maximum likelihood (ML) methods, apparently due to their amino acid sequence diversity. Only a few internal clades in the ML tree showed bootstrap support values higher than 50%. The phylogram in Figure 4A reveals five well-supported clades of Kunitz protease inhibitors in soft and hard ticks, scorpions and horse flies.

Bottom Line: Using homology-based modeling (and other protein prediction programs) we were able to model and explain the multifaceted function of tryptogalinin.The N-terminus of the modeled tryptogalinin is detached from the rest of the peptide and exhibits intrinsic disorder allowing an increased flexibility for its high affinity with its inhibiting partners (i.e., serine proteases).By incorporating experimental and computational methods our data not only describes the function of a Kunitz peptide from Ixodes scapularis, but also allows us to hypothesize about the molecular basis of this function at the atomic level.

View Article: PubMed Central - PubMed

Affiliation: Institute of Parasitology, Biology Centre of the Academy of Sciences of the Czech Republic, České Budějovice, Czech Republic. valdjj@gmail.com

ABSTRACT

Background: A salivary proteome-transcriptome project on the hard tick Ixodes scapularis revealed that Kunitz peptides are the most abundant salivary proteins. Ticks use Kunitz peptides (among other salivary proteins) to combat host defense mechanisms and to obtain a blood meal. Most of these Kunitz peptides, however, remain functionally uncharacterized, thus limiting our knowledge about their biochemical interactions.

Results: We discovered an unusual cysteine motif in a Kunitz peptide. This peptide inhibits several serine proteases with high affinity and was named tryptogalinin due to its high affinity for β-tryptase. Compared with other functionally described peptides from the Acari subclass, we showed that tryptogalinin is phylogenetically related to a Kunitz peptide from Rhipicephalus appendiculatus, also reported to have a high affinity for β-tryptase. Using homology-based modeling (and other protein prediction programs) we were able to model and explain the multifaceted function of tryptogalinin. The N-terminus of the modeled tryptogalinin is detached from the rest of the peptide and exhibits intrinsic disorder allowing an increased flexibility for its high affinity with its inhibiting partners (i.e., serine proteases).

Conclusions: By incorporating experimental and computational methods our data not only describes the function of a Kunitz peptide from Ixodes scapularis, but also allows us to hypothesize about the molecular basis of this function at the atomic level.

Show MeSH