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Diaz Perales, AraceliAuthor

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June 9, 2019
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Article

Purification and Characterization of AsES Protein A SUBTILISIN SECRETED BY ACREMONIUM STRICTUM IS A NOVEL PLANT DEFENSE ELICITOR

Publicated to: JOURNAL OF BIOLOGICAL CHEMISTRY. 288 (20): 14098-14113 - 2013-05-17 288(20), DOI: 10.1074/jbc.M112.429423

Authors:

Chalfoun, NR; Grellet-Bournonville, CF; Martínez-Zamora, MG; Díaz-Perales, A; Castagnaro, AP; Díaz-Ricci, JC
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Affiliations

Estn Expt Agroind Obispo Colombres, Secc Biotecnol, Las Talitas - Author
Univ Nacl Tucuman, Fac Bioquim Quim & Farm, Consejo Nacl Invest Cient & Tecn, RA-4000 San Miguel De Tucuman - Author
Univ Politecn Madrid, Escuela Tecn Super Ingenieros Agronomos, Dept Biotecnol, Unidad Quim & Bioquim - Author
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Abstract

In this work, the purification and characterization of an extracellular elicitor protein, designated AsES, produced by an avirulent isolate of the strawberry pathogen Acremonium strictum, are reported. The defense eliciting activity present in culture filtrates was recovered and purified by ultrafiltration (cutoff, 30 kDa), anionic exchange (Q-Sepharose, pH 7.5), and hydrophobic interaction (phenyl-Sepharose) chromatographies. Two-dimensional SDS-PAGE of the purified active fraction revealed a single spot of 34 kDa and pI 8.8. HPLC (C2/C18) and MS/MS analysis confirmed purification to homogeneity. Foliar spray with AsES provided a total systemic protection against anthracnose disease in strawberry, accompanied by the expression of defense-related genes (i.e. PR1 and Chi2-1). Accumulation of reactive oxygen species (e.g. H2O2 and O-2(radical anion)) and callose was also observed in Arabidopsis. By using degenerate primers designed from the partial amino acid sequences and rapid amplification reactions of cDNA ends, the complete AsES-coding cDNA of 1167 nucleotides was obtained. The deduced amino acid sequence showed significant identity with fungal serine proteinases of the subtilisin family, indicating that AsES is synthesized as a larger precursor containing a 15-residue secretory signal peptide and a 90-residue peptidase inhibitor I9 domain in addition to the 283-residue mature protein. AsES exhibited proteolytic activity in vitro, and its resistance eliciting activity was eliminated when inhibited with PMSF, suggesting that its proteolytic activity is required to induce the defense response. This is, to our knowledge, the first report of a fungal subtilisin that shows eliciting activity in plants. This finding could contribute to develop disease biocontrol strategies in plants by activating its innate immunity.
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Keywords

acremonium strictumelicitorfragaria x ananassamass spectrometry (ms)plant defenseprotein purificationprotein sequenceserine proteasesubtilisinAcremoniumAcremonium strictumAmino acid sequenceArabidopsisAvirulent strainBase sequenceBiological assayCdna sequencingChromatography, high pressure liquidCloning, molecularDisease resistanceDna, complementaryElectrophoresis, polyacrylamide gelElicitorEscherichia-coliFragariaFragaria x ananassaFungal effector proteinsFungal proteinsHypersensitive responseMass spectrometryMass spectrometry (ms)Mass-spectrometryMolecular sequence dataNucleotide-sequencesPathogenPlant defensePlant diseasesPlant immunityProtein purificationProtein sequenceReactive oxygen speciesResistanceSequence analysis, dnaSerine proteaseSerine proteasesSpectrometry, mass, matrix-assisted laser desorption-ionizationStrawberrySubtilisinSubtilisinsTrypsinUltrafiltration

Quality index

Bibliometric impact. Analysis of the contribution and dissemination channel

The work has been published in the journal JOURNAL OF BIOLOGICAL CHEMISTRY due to its progression and the good impact it has achieved in recent years, according to the agency WoS (JCR), it has become a reference in its field. In the year of publication of the work, 2013, it was in position 65/291, thus managing to position itself as a Q1 (Primer Cuartil), in the category Biochemistry & Molecular Biology.

From a relative perspective, and based on the normalized impact indicator calculated from World Citations from Scopus Elsevier, it yields a value for the Field-Weighted Citation Impact from the Scopus agency: 1.11, which indicates that, compared to works in the same discipline and in the same year of publication, it ranks as a work cited above average. (source consulted: ESI Nov 13, 2025)

Specifically, and according to different indexing agencies, this work has accumulated citations as of 2026-04-24, the following number of citations:

  • WoS: 45
  • Scopus: 47
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Impact and social visibility

From the perspective of influence or social adoption, and based on metrics associated with mentions and interactions provided by agencies specializing in calculating the so-called "Alternative or Social Metrics," we can highlight as of 2026-04-24:

  • The use, from an academic perspective evidenced by the Altmetric agency indicator referring to aggregations made by the personal bibliographic manager Mendeley, gives us a total of: 57.
  • The use of this contribution in bookmarks, code forks, additions to favorite lists for recurrent reading, as well as general views, indicates that someone is using the publication as a basis for their current work. This may be a notable indicator of future more formal and academic citations. This claim is supported by the result of the "Capture" indicator, which yields a total of: 64 (PlumX).

With a more dissemination-oriented intent and targeting more general audiences, we can observe other more global scores such as:

  • The Total Score from Altmetric: 10.
  • The number of mentions on the social network X (formerly Twitter): 1 (Altmetric).
  • The number of mentions on Wikipedia: 2 (Altmetric).

It is essential to present evidence supporting full alignment with institutional principles and guidelines on Open Science and the Conservation and Dissemination of Intellectual Heritage. A clear example of this is:

  • The work has been submitted to a journal whose editorial policy allows open Open Access publication.
  • Assignment of a Handle/URN as an identifier within the deposit in the Institutional Repository: https://oa.upm.es/26288/

As a result of the publication of the work in the institutional repository, statistical usage data has been obtained that reflects its impact. In terms of dissemination, we can state that, as of

  • Views: 403
  • Downloads: 228
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Leadership analysis of institutional authors

This work has been carried out with international collaboration, specifically with researchers from: Argentina.

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Awards linked to the item

This work was supported in part by Consejo de Investigaciones de la Universidad Nacional de Tucuman Grant 26/D423 and Agencia Nacional de Promocion Cientifica y Tecnologica Grant BID PICT-2008-2105.
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