Identification and recombinant expression of an antimicrobial peptide (cecropin B-like) from soybean pest Anticarsia gemmatalis
Abstract Background Insects can be found in numerous diverse environments, being exposed to pathogenic organisms like fungi and bacteria. Once these pathogens cross insect physical barriers, the innate immune system operates through cellular and humoral responses. Antimicrobial peptides are small m...
| Main Authors: | , , , , , , , , , , |
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| Format: | Article |
| Language: | English |
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SciELO
2021-03-01
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| Series: | Journal of Venomous Animals and Toxins including Tropical Diseases |
| Subjects: | |
| Online Access: | http://www.scielo.br/scielo.php?script=sci_arttext&pid=S1678-91992021000100305&tlng=en |
| _version_ | 1818999691213275136 |
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| author | Luís Felipe Costa Ramos João Henrique de Oliveira Rangel Guilherme Caldas Andrade Carolina Lixa Livia Vieira Araujo de Castilho Fábio César Sousa Nogueira Anderson S. Pinheiro Fabio Mendonça Gomes Cristiane Dinis AnoBom Rodrigo Volcan Almeida Danielle Maria Perpétua de Oliveira |
| author_facet | Luís Felipe Costa Ramos João Henrique de Oliveira Rangel Guilherme Caldas Andrade Carolina Lixa Livia Vieira Araujo de Castilho Fábio César Sousa Nogueira Anderson S. Pinheiro Fabio Mendonça Gomes Cristiane Dinis AnoBom Rodrigo Volcan Almeida Danielle Maria Perpétua de Oliveira |
| author_sort | Luís Felipe Costa Ramos |
| collection | DOAJ |
| description | Abstract Background Insects can be found in numerous diverse environments, being exposed to pathogenic organisms like fungi and bacteria. Once these pathogens cross insect physical barriers, the innate immune system operates through cellular and humoral responses. Antimicrobial peptides are small molecules produced by immune signaling cascades that develop an important and generalist role in insect defenses against a variety of microorganisms. In the present work, a cecropin B-like peptide (AgCecropB) sequence was identified in the velvetbean caterpillar Anticarsia gemmatalis and cloned in a bacterial plasmid vector for further heterologous expression and antimicrobial tests. Methods AgCecropB sequence (without the signal peptide) was cloned in the plasmid vector pET-M30-MBP and expressed in the Escherichia coli BL21(DE3) expression host. Expression was induced with IPTG and a recombinant peptide was purified using two affinity chromatography steps with Histrap column. The purified peptide was submitted to high-resolution mass spectrometry (HRMS) and structural analyses. Antimicrobial tests were performed using gram-positive (Bacillus thuringiensis) and gram-negative (Burkholderia kururiensis and E. coli) bacteria. Results AgCecropB was expressed in E. coli BL21 (DE3) at 28°C with IPTG 0.5 mM. The recombinant peptide was purified and enriched after purification steps. HRMS confirmed AgCrecropB molecular mass (4.6 kDa) and circular dichroism assay showed α-helix structure in the presence of SDS. AgCrecropB inhibited almost 50% of gram-positive B. thuringiensis bacteria growth. Conclusions The first cecropin B-like peptide was described in A. gemmatalis and a recombinant peptide was expressed using a bacterial platform. Data confirmed tertiary structure as predicted for the cecropin peptide family. AgCecropB was capable to inhibit B. thuringiensis growth in vitro. |
| first_indexed | 2024-12-20T22:21:27Z |
| format | Article |
| id | doaj.art-4e48b8817435421fa18004621d545228 |
| institution | Directory Open Access Journal |
| issn | 1678-9199 |
| language | English |
| last_indexed | 2024-12-20T22:21:27Z |
| publishDate | 2021-03-01 |
| publisher | SciELO |
| record_format | Article |
| series | Journal of Venomous Animals and Toxins including Tropical Diseases |
| spelling | doaj.art-4e48b8817435421fa18004621d5452282022-12-21T19:24:56ZengSciELOJournal of Venomous Animals and Toxins including Tropical Diseases1678-91992021-03-012710.1590/1678-9199-jvatitd-2020-0127Identification and recombinant expression of an antimicrobial peptide (cecropin B-like) from soybean pest Anticarsia gemmatalisLuís Felipe Costa RamosJoão Henrique de Oliveira RangelGuilherme Caldas AndradeCarolina LixaLivia Vieira Araujo de CastilhoFábio César Sousa NogueiraAnderson S. PinheiroFabio Mendonça GomesCristiane Dinis AnoBomRodrigo Volcan Almeidahttps://orcid.org/0000-0002-0164-6044Danielle Maria Perpétua de Oliveirahttps://orcid.org/0000-0002-9889-5556Abstract Background Insects can be found in numerous diverse environments, being exposed to pathogenic organisms like fungi and bacteria. Once these pathogens cross insect physical barriers, the innate immune system operates through cellular and humoral responses. Antimicrobial peptides are small molecules produced by immune signaling cascades that develop an important and generalist role in insect defenses against a variety of microorganisms. In the present work, a cecropin B-like peptide (AgCecropB) sequence was identified in the velvetbean caterpillar Anticarsia gemmatalis and cloned in a bacterial plasmid vector for further heterologous expression and antimicrobial tests. Methods AgCecropB sequence (without the signal peptide) was cloned in the plasmid vector pET-M30-MBP and expressed in the Escherichia coli BL21(DE3) expression host. Expression was induced with IPTG and a recombinant peptide was purified using two affinity chromatography steps with Histrap column. The purified peptide was submitted to high-resolution mass spectrometry (HRMS) and structural analyses. Antimicrobial tests were performed using gram-positive (Bacillus thuringiensis) and gram-negative (Burkholderia kururiensis and E. coli) bacteria. Results AgCecropB was expressed in E. coli BL21 (DE3) at 28°C with IPTG 0.5 mM. The recombinant peptide was purified and enriched after purification steps. HRMS confirmed AgCrecropB molecular mass (4.6 kDa) and circular dichroism assay showed α-helix structure in the presence of SDS. AgCrecropB inhibited almost 50% of gram-positive B. thuringiensis bacteria growth. Conclusions The first cecropin B-like peptide was described in A. gemmatalis and a recombinant peptide was expressed using a bacterial platform. Data confirmed tertiary structure as predicted for the cecropin peptide family. AgCecropB was capable to inhibit B. thuringiensis growth in vitro.http://www.scielo.br/scielo.php?script=sci_arttext&pid=S1678-91992021000100305&tlng=enAntimicrobial peptidesCecropin BHeterologous expressionAnticarsia gemmatalisAgricultural pest |
| spellingShingle | Luís Felipe Costa Ramos João Henrique de Oliveira Rangel Guilherme Caldas Andrade Carolina Lixa Livia Vieira Araujo de Castilho Fábio César Sousa Nogueira Anderson S. Pinheiro Fabio Mendonça Gomes Cristiane Dinis AnoBom Rodrigo Volcan Almeida Danielle Maria Perpétua de Oliveira Identification and recombinant expression of an antimicrobial peptide (cecropin B-like) from soybean pest Anticarsia gemmatalis Journal of Venomous Animals and Toxins including Tropical Diseases Antimicrobial peptides Cecropin B Heterologous expression Anticarsia gemmatalis Agricultural pest |
| title | Identification and recombinant expression of an antimicrobial peptide (cecropin B-like) from soybean pest Anticarsia gemmatalis |
| title_full | Identification and recombinant expression of an antimicrobial peptide (cecropin B-like) from soybean pest Anticarsia gemmatalis |
| title_fullStr | Identification and recombinant expression of an antimicrobial peptide (cecropin B-like) from soybean pest Anticarsia gemmatalis |
| title_full_unstemmed | Identification and recombinant expression of an antimicrobial peptide (cecropin B-like) from soybean pest Anticarsia gemmatalis |
| title_short | Identification and recombinant expression of an antimicrobial peptide (cecropin B-like) from soybean pest Anticarsia gemmatalis |
| title_sort | identification and recombinant expression of an antimicrobial peptide cecropin b like from soybean pest anticarsia gemmatalis |
| topic | Antimicrobial peptides Cecropin B Heterologous expression Anticarsia gemmatalis Agricultural pest |
| url | http://www.scielo.br/scielo.php?script=sci_arttext&pid=S1678-91992021000100305&tlng=en |
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