Lytic Activity and Structural Differences of Amphipathic Peptides Derived from Trialysin^,

Abstract

Trialysin is a pore-forming protein found in the saliva of Triatoma infestans (Hemiptera, Reduviidae), the insect vector of Chagas' disease. The protein is active against a broad range of cell types from bacteria to eukaryotic cells. Recognizing that the N-terminus of trialysin harbors the lytic motif [Amino, R., Martins, R. M., Procopio, J., Hirata, I. Y., Juliano, M. A., and Schenkman, S. (2002) J. Biol. Chem. 277, 6207-6213], we designed a set of peptides scanning this region to investigate the structural basis of its biological function. Peptides encompassing residues 1-32 (P6), 1-27 (P7), and 6-32 (P5) efficiently induced lysis of the protozoan parasite Trypanosoma cruzi and Escherichia coli in the 0.4-9.0 microM range, while much higher concentrations were required to cause hemolysis. Other more internal peptides, including peptide P2 (residues 21-47) and others up to residue 52, were less effective. P6 turned out to be the most active of all. P7 has a significantly higher activity than P5 against E. coli, while P5 has a hemolytic activity comparable to that of P6. CD spectroscopy showed that all tested peptides acquire a comparable helical content in solvent mixtures or in detergent micelles. The solution structure of P2 and P5-P7 was determined in a 30% trifluoroethanol/water mixture by nuclear magnetic resonance. All peptides exhibit a structure characterized by a central helical fold, and except for P2, which does not show a continuous hydrophobic surface, they are amphipathic. The structural models show that P5 and P7 extend their structural similarities with the most active peptide, P6, in either the C-terminus or the N-terminus. Amino acid substitutions in the N-terminus of P6 improved hemolysis but did not change the activity against T. cruzi. These results suggest that while amphipathicity is essential for the lytic activity, the selectivity of the active peptides for specific organisms appears to be associated with the structural features of their N- and C-termini.

An insight into the sialome of the blood-sucking bug Triatoma infestans, a vector of Chagas’ disease

“…3.5.2 Coding for proteins found only in triatomines: 3.5.2.1 Trialysin: We identified two contigs that matched the protein trialysin, a pore-forming molecule present in the (SG)s of T. infestans Martins et al, 2006) (Fig. 9).…”

“…Studies in which synthetic peptides containing portions of the Nterminus of trialysin have been tested for their lytic properties against the infective stage of T. cruzi, Escherichia coli and human erythrocytes revealed differences in specificity and activity for the tested targets. The cell membrane is the main target of antimicrobial peptides, thus the diverse lipid composition and distribution or the presence of other components in the cell membrane of the various organisms play an important role in the modulation of peptide/ membrane interaction, leading to the differentiation of their action Martins et al, 2006).…”

An Insight into the Sialotranscriptome of Triatoma matogrossensis, a Kissing Bug Associated with Fogo Selvagem in South America

“…We identified six contigs (assembled from 54 ESTs that matched the protein named trialysin from T. infestans 77,78 ) having between 38% and 88% identity at the aa level with their best match in the NR database, all from T. infestans. Trialysin, which belongs to a multigene family, is a pore-forming molecule present in the SGs of T. infestans.…”

“…The structural features of the protein and charge distribution, as well as the composition of the target membrane, are important determinants for protein/membrane interaction and lysis specificity. 77,78 Antigen 5 family. This family of secreted proteins belongs to the CAP family (cysteine-rich secretory proteins; Ag5 proteins of insects; pathogenesis-related protein 1 of plants).…”

The structural molecular biology network of the State of São Paulo, Brazil

“…In parallel, we studied the lytic activity of several synthetic peptides corresponding to the Nterminus of trialysin (Martins et al 2006). We found that although all peptides folded into α-helices in the presence of SDS or trifluoroethanol (TFE), only the ones close to the N-terminus of mature trialysin were lytic to T. cruzi, E. coli, and erythrocytes.…”