In-depth characterization of trypsin-like serine peptidases in the midgut of the sugar fed Culex quinquefasciatus
Abstract Background Culex quinquefasciatus is a hematophagous insect from the Culicidae family that feeds on the blood of humans, dogs, birds and livestock. This species transmits a wide variety of pathogens between humans and animals. The midgut environment is the first location of pathogen-vector...
| Main Authors: | , , , , , , |
|---|---|
| Format: | Article |
| Language: | English |
| Published: |
BMC
2015-07-01
|
| Series: | Parasites & Vectors |
| Subjects: | |
| Online Access: | https://doi.org/10.1186/s13071-015-0985-0 |
| _version_ | 1797811840844038144 |
|---|---|
| author | André Borges-Veloso Leonardo Saboia-Vahia Geovane Dias-Lopes Gilberto B. Domont Constança Britto Patricia Cuervo Jose B. De Jesus |
| author_facet | André Borges-Veloso Leonardo Saboia-Vahia Geovane Dias-Lopes Gilberto B. Domont Constança Britto Patricia Cuervo Jose B. De Jesus |
| author_sort | André Borges-Veloso |
| collection | DOAJ |
| description | Abstract Background Culex quinquefasciatus is a hematophagous insect from the Culicidae family that feeds on the blood of humans, dogs, birds and livestock. This species transmits a wide variety of pathogens between humans and animals. The midgut environment is the first location of pathogen-vector interactions for blood-feeding mosquitoes and the expression of specific peptidases in the early stages of feeding could influence the outcome of the infection. Trypsin-like serine peptidases belong to a multi-gene family that can be expressed in different isoforms under distinct physiological conditions. However, the confident assignment of the trypsin genes that are expressed under each condition is still a challenge due to the large number of trypsin-coding genes in the Culicidae family and most likely because they are low abundance proteins. Methods We used zymography for the biochemical characterization of the peptidase profile of the midgut from C. quinquefasciatus females fed on sugar. Protein samples were also submitted to SDS-PAGE followed by liquid chromatography–tandem mass spectrometry (LC–MS/MS) analysis for peptidase identification. The peptidases sequences were analyzed with bioinformatics tools to assess their distinct features. Results Zymography revealed that trypsin-like serine peptidases were responsible for the proteolytic activity in the midgut of females fed on sugar diet. After denaturation in SDS-PAGE, eight trypsin-like serine peptidases were identified by LC-MS/MS. These peptidases have structural features typical of invertebrate digestive trypsin peptidases but exhibited singularities at the protein sequence level such as: the presence of different amino acids at the autocatalytic motif and substrate binding regions as well as different number of disulfide bounds. Data mining revealed a group of trypsin-like serine peptidases that are specific to C. quinquefasciatus when compared to the culicids genomes sequenced so far. Conclusion We demonstrated that proteomics approaches combined with bioinformatics tools and zymographic analysis can lead to the functional annotation of trypsin-like serine peptidases coding genes and aid in the understanding of the complexity of peptidase expression in mosquitoes. |
| first_indexed | 2024-03-13T07:29:45Z |
| format | Article |
| id | doaj.art-b7b84099c62948cda3cf1067e51f7db3 |
| institution | Directory Open Access Journal |
| issn | 1756-3305 |
| language | English |
| last_indexed | 2024-03-13T07:29:45Z |
| publishDate | 2015-07-01 |
| publisher | BMC |
| record_format | Article |
| series | Parasites & Vectors |
| spelling | doaj.art-b7b84099c62948cda3cf1067e51f7db32023-06-04T11:12:29ZengBMCParasites & Vectors1756-33052015-07-018111610.1186/s13071-015-0985-0In-depth characterization of trypsin-like serine peptidases in the midgut of the sugar fed Culex quinquefasciatusAndré Borges-Veloso0Leonardo Saboia-Vahia1Geovane Dias-Lopes2Gilberto B. Domont3Constança Britto4Patricia Cuervo5Jose B. De Jesus6Laboratório de Biologia Molecular e Doenças Endêmicas, Instituto Oswaldo Cruz, FIOCRUZLaboratorio de Pesquisa em Leishmaniose, Instituto Oswaldo Cruz, FIOCRUZLaboratório de Biologia Molecular e Doenças Endêmicas, Instituto Oswaldo Cruz, FIOCRUZUnidade de Proteômica, Laboratório de Química de Proteínas, Instituto de Química, Universidade Federal do Rio de JaneiroLaboratório de Biologia Molecular e Doenças Endêmicas, Instituto Oswaldo Cruz, FIOCRUZLaboratorio de Pesquisa em Leishmaniose, Instituto Oswaldo Cruz, FIOCRUZLaboratório de Biologia Molecular e Doenças Endêmicas, Instituto Oswaldo Cruz, FIOCRUZAbstract Background Culex quinquefasciatus is a hematophagous insect from the Culicidae family that feeds on the blood of humans, dogs, birds and livestock. This species transmits a wide variety of pathogens between humans and animals. The midgut environment is the first location of pathogen-vector interactions for blood-feeding mosquitoes and the expression of specific peptidases in the early stages of feeding could influence the outcome of the infection. Trypsin-like serine peptidases belong to a multi-gene family that can be expressed in different isoforms under distinct physiological conditions. However, the confident assignment of the trypsin genes that are expressed under each condition is still a challenge due to the large number of trypsin-coding genes in the Culicidae family and most likely because they are low abundance proteins. Methods We used zymography for the biochemical characterization of the peptidase profile of the midgut from C. quinquefasciatus females fed on sugar. Protein samples were also submitted to SDS-PAGE followed by liquid chromatography–tandem mass spectrometry (LC–MS/MS) analysis for peptidase identification. The peptidases sequences were analyzed with bioinformatics tools to assess their distinct features. Results Zymography revealed that trypsin-like serine peptidases were responsible for the proteolytic activity in the midgut of females fed on sugar diet. After denaturation in SDS-PAGE, eight trypsin-like serine peptidases were identified by LC-MS/MS. These peptidases have structural features typical of invertebrate digestive trypsin peptidases but exhibited singularities at the protein sequence level such as: the presence of different amino acids at the autocatalytic motif and substrate binding regions as well as different number of disulfide bounds. Data mining revealed a group of trypsin-like serine peptidases that are specific to C. quinquefasciatus when compared to the culicids genomes sequenced so far. Conclusion We demonstrated that proteomics approaches combined with bioinformatics tools and zymographic analysis can lead to the functional annotation of trypsin-like serine peptidases coding genes and aid in the understanding of the complexity of peptidase expression in mosquitoes.https://doi.org/10.1186/s13071-015-0985-0Culex quinquefasciatusTrypsin-like serine peptidasesZymographyMass spectrometry |
| spellingShingle | André Borges-Veloso Leonardo Saboia-Vahia Geovane Dias-Lopes Gilberto B. Domont Constança Britto Patricia Cuervo Jose B. De Jesus In-depth characterization of trypsin-like serine peptidases in the midgut of the sugar fed Culex quinquefasciatus Parasites & Vectors Culex quinquefasciatus Trypsin-like serine peptidases Zymography Mass spectrometry |
| title | In-depth characterization of trypsin-like serine peptidases in the midgut of the sugar fed Culex quinquefasciatus |
| title_full | In-depth characterization of trypsin-like serine peptidases in the midgut of the sugar fed Culex quinquefasciatus |
| title_fullStr | In-depth characterization of trypsin-like serine peptidases in the midgut of the sugar fed Culex quinquefasciatus |
| title_full_unstemmed | In-depth characterization of trypsin-like serine peptidases in the midgut of the sugar fed Culex quinquefasciatus |
| title_short | In-depth characterization of trypsin-like serine peptidases in the midgut of the sugar fed Culex quinquefasciatus |
| title_sort | in depth characterization of trypsin like serine peptidases in the midgut of the sugar fed culex quinquefasciatus |
| topic | Culex quinquefasciatus Trypsin-like serine peptidases Zymography Mass spectrometry |
| url | https://doi.org/10.1186/s13071-015-0985-0 |
| work_keys_str_mv | AT andreborgesveloso indepthcharacterizationoftrypsinlikeserinepeptidasesinthemidgutofthesugarfedculexquinquefasciatus AT leonardosaboiavahia indepthcharacterizationoftrypsinlikeserinepeptidasesinthemidgutofthesugarfedculexquinquefasciatus AT geovanediaslopes indepthcharacterizationoftrypsinlikeserinepeptidasesinthemidgutofthesugarfedculexquinquefasciatus AT gilbertobdomont indepthcharacterizationoftrypsinlikeserinepeptidasesinthemidgutofthesugarfedculexquinquefasciatus AT constancabritto indepthcharacterizationoftrypsinlikeserinepeptidasesinthemidgutofthesugarfedculexquinquefasciatus AT patriciacuervo indepthcharacterizationoftrypsinlikeserinepeptidasesinthemidgutofthesugarfedculexquinquefasciatus AT josebdejesus indepthcharacterizationoftrypsinlikeserinepeptidasesinthemidgutofthesugarfedculexquinquefasciatus |