Three-dimensional crystal structure of human eosinophil cationic protein (RNase 3) at 1.75 Å resolution11Edited by R. Huber

Mallorqui-Fernandez, G.; Pous, Joan; Peracaula, Rosa; Aymamı́, Joan; Maeda, Takashi; Tada, Hayato; Yamada, Hidenori; Seno, Masaharu; Llorens, Rafael de; Gomis‐Rüth, F. Xavier; Coll, Miquel

doi:10.1006/jmbi.2000.3939

Cited by 50 publications

(57 citation statements)

References 54 publications

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“…Residue W35, exposed to the aqueous solvent at the ECP protein surface, would find itself in an unfavorable position and may tend to bury itself by interacting with a lipid bilayer. A close look at the ECP crystallographic dimer reported by ref 35 shows that W35 is located on the interacting interfaces of two ECP molecules. The dimer interface surface includes W35 together with discrete hydrophobic patches at the protein N-terminus and C-terminal loop.…”

Section: Discussionmentioning

confidence: 75%

“…The data are summarized in Table 3. The two molecules of the ECP crystallographic dimer structure [PDB entry code 1DYT (35)] show some differences on the structural parameters for the W10 residue microenvironment. In fact, molecular dynamics simulation studies on the ECP crystallographic dimer (46) indicate that main significant differences between the two units lie mainly at the ECP N terminus.…”

Section: Protein Fluorescence Properties In the Absence And Presence mentioning

confidence: 99%

“…Trp residues were classified and assigned to distinct classes using an algorithm as described in ref 31. b PDB entry code 1QMT (34). c The mean values of chains A and B. d PDB entry code 1DYT (35). e PDB entry code 1H1H (36).…”

Section: Depth-dependent Fluorescence Quenching Experiments Using Labmentioning

confidence: 99%

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Topography Studies on the Membrane Interaction Mechanism of the Eosinophil Cationic Protein

et al. 2006

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The eosinophil cationic protein (ECP) is an antipathogen protein involved in the host defense system. ECP displays bactericidal and membrane lytic capacities [Carreras et al. (2003) Biochemistry 42, 6636-6644]. We have now characterized in detail the protein-membrane interaction process. All observed fluorescent parameters of the wild type and single-tryptophan-containing mutants, as well as the results of decomposition analysis of protein fluorescence, suggest that W10 and W35 belong to two distinct spectral classes I and III, respectively. Tryptophan residues were classified and assigned to distinct structural classes using statistical approaches based on the analysis of tryptophan microenvironment structural properties. W10 belongs to class I and is buried in a relative nonpolar, nonflexible protein environment, while W35 (class III) is fully exposed to free water molecules. Tryptophan solvent exposure and the depth of the protein insertion in the lipid bilayer were monitored by the degree of protein fluorescence quenching by KI and brominated phospholipids, respectively. Results indicate that W35 partially inserts into the lipid bilayer, whereas W10 does not. Further analysis by electron microscopy and dynamic light scattering indicates that ECP can destabilize and trigger lipid vesicle aggregation at a nanomolar concentration range, corresponding to about 1:1000 protein/lipid ratio. No significant leakage of the vesicle aqueous content takes place below that protein concentration threshold. The data are consistent with a membrane destabilization "carpet-like" mechanism.

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Section: Discussionmentioning

confidence: 75%

Section: Protein Fluorescence Properties In the Absence And Presence mentioning

confidence: 99%

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Topography Studies on the Membrane Interaction Mechanism of the Eosinophil Cationic Protein

et al. 2006

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“…La Jolla, CA). ECP structure 1DYT.pdb (22) was used as receptor molecule, and LPS ligand was obtained from 1FI1.pdb (13). Water molecules were removed from the structure; hydrogen atoms and atomic partial charges were added using Autodock Tools.…”

Section: Methodsmentioning

confidence: 99%

Antimicrobial Action and Cell Agglutination by the Eosinophil Cationic Protein Are Modulated by the Cell Wall Lipopolysaccharide Structure

Pulido

Moussaoui

Andreu

et al. 2012

Antimicrob Agents Chemother

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Antimicrobial proteins and peptides (AMPs) are essential effectors of innate immunity, acting as a first line of defense against bacterial infections. Many AMPs exhibit high affinity for cell wall structures such as lipopolysaccharide (LPS), a potent endotoxin able to induce sepsis. Hence, understanding how AMPs can interact with and neutralize LPS endotoxin is of special relevance for human health. Eosinophil cationic protein (ECP) is an eosinophil secreted protein with high activity against both Gram-negative and Gram-positive bacteria. ECP has a remarkable affinity for LPS and a distinctive agglutinating activity. By using a battery of LPS-truncated E. coli mutant strains, we demonstrate that the polysaccharide moiety of LPS is essential for ECP-mediated bacterial agglutination, thereby modulating its antimicrobial action. The mechanism of action of ECP at the bacterial surface is drastically affected by the LPS structure and in particular by its polysaccharide moiety. We have also analyzed an N-terminal fragment that retains the whole protein activity and displays similar cell agglutination behavior. Conversely, a fragment with further minimization of the antimicrobial domain, though retaining the antimicrobial capacity, significantly loses its agglutinating activity, exhibiting a different mechanism of action which is not dependent on the LPS composition. The results highlight the correlation between the protein's antimicrobial activity and its ability to interact with the LPS outer layer and promote bacterial agglutination.

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“…9 Briefly, human ECP cDNA was isolated and expressed in an Escherichia coli T7 expression system. Purified rhECP was assessed for RNase activity on yeast RNA by the perchloric acid precipitation method 10 and for bactericidal activity.…”

Section: Recombinant Human Ecpmentioning

confidence: 99%

Cytokine expression in human dermal fibroblasts stimulated with eosinophil cationic protein measured by protein array

Sato

Soga²,

Yamaguchi

et al. 2013

Asian Pac J Allergy Immunol

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Summa ryBackground: Eosinophil cationic protein (ECP) was reported previously to be involved in allergic inflammation with cytotoxic activity. On the other hand, recent studies showed that ECP did not induce cell death but inhibited the growth of cancer-derived cells. Our previous study indicated that human ECP enhanced differentiation of rat neonatal cardiomyocytes and stress fiber formation in Balb/c 3T3 mouse fibroblasts, while the effects of human ECP on human fibroblasts are unknown.Objective: The present study was performed to determine the effects of human ECP on cytokine expression in human fibroblasts by protein array. Methods:The effects of recombinant human ECP (rhECP) on normal human dermal fibroblasts (NHDF) were examined by assaying cell growth. Furthermore, cytokine expression of NHDF stimulated by ECP, which could influence cell growth, was evaluated by protein array.Results: ECP was not cytotoxic but enhanced the growth of NHDF. The peak rhECP concentration that enhanced the cell counts by 1.56-fold was 100 ng/mL, which was significantly different from cultures without ECP stimulation (ANOVA/ Scheffe's test, P < 0.05). Array analyses indicated that ciliary neurotrophic factor (CNTF), neutrophilactivating peptide (NAP)-2, and neurotrophin (NT)-3 were significantly upregulated in NHDF stimulated with 100 ng/mL ECP compared to those without stimulation. Conclusion

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Three-dimensional crystal structure of human eosinophil cationic protein (RNase 3) at 1.75 Å resolution11Edited by R. Huber

Cited by 50 publications

References 54 publications

Topography Studies on the Membrane Interaction Mechanism of the Eosinophil Cationic Protein

Topography Studies on the Membrane Interaction Mechanism of the Eosinophil Cationic Protein

Antimicrobial Action and Cell Agglutination by the Eosinophil Cationic Protein Are Modulated by the Cell Wall Lipopolysaccharide Structure

Cytokine expression in human dermal fibroblasts stimulated with eosinophil cationic protein measured by protein array

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