{"title":"丝光Lucilia sericata幼虫唾液腺Netrin-A的鉴定、分子特征和硅结构分析","authors":"Masoumeh Bagheri, Hamzeh Alipour, Tahereh Karamzadeh, Marzieh Shahriari-Namadi, Abbasali Raz, Kourosh Azizi, Javad Dadgar Pakdel, Mohammad Djaefar Moemenbellah-Fard","doi":"10.1007/s10709-022-00164-8","DOIUrl":null,"url":null,"abstract":"<p><p>The greenbottle blowfly Lucilia sericata (L. sericata) is increasingly used in larval therapy of chronic wounds. Netrins as bifunctional proteins are in the superfamily of Laminins secreted from larval salivary glands. The Netrin protein has a significant instructive role in axon guidance, causing neuronal outgrowth, angiogenesis, and cell migration. It seems to be crucial in wound healing and acts as a potential biomarker in diagnosing some clinical diseases. This survey aimed to identify molecular features and analyze in silico structural configuration of Netrin-A in L. sericata larvae. The larvae were reared under standard maggotarium conditions. The nucleic acid sequence of L. sericata Netrin-A (LSN-A) was then identified using rapid amplification of circular DNA ends (RACE) and rapid amplification of genomic ends (RAGE). Parts of the Netrin-A gene, including the middle, 3'-, and 5'-ends, were identified, TA cloned in pTG19 plasmid, and transferred into DH5ɑ Escherichia coli. Each part was sequenced and assembled using SeqMan software. This gene structure was further subjected to in silico analysis. The DNA of LSN-A was identified to be 2407 bp, while its mRNA sequence was recognized as 2115 bp by Oligo0.7 software. It translated the Netrin-A protein with 704 amino acid residues. Its estimated molecular weight was 78.6 kDa. Sequencing of this fragment and its BLAST analysis revealed laminin-based high (95%) similarity with the mRNA sequence of Lucilia cuprina Netrin-A. The 3-D structure of Netrin-A drawn by SWISS-MODEL exhibited its partial resemblance to the reference molecule Netrin-1 of Homo sapiens. This study supports the molecular and structural analyses of LSN-A protein, which could lead to wound treatment. Ultimately, it can be an effective candidate to ameliorate injury. Our next attempt is to produce LSN-A recombinant protein for use in biomedical sciences.</p>","PeriodicalId":1,"journal":{"name":"Accounts of Chemical Research","volume":null,"pages":null},"PeriodicalIF":16.4000,"publicationDate":"2022-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":"{\"title\":\"Identification, molecular characterization, and in silico structural analysis of larval salivary glands Netrin-A as a potent biomarker from Lucilia sericata (Diptera: Calliphoridae).\",\"authors\":\"Masoumeh Bagheri, Hamzeh Alipour, Tahereh Karamzadeh, Marzieh Shahriari-Namadi, Abbasali Raz, Kourosh Azizi, Javad Dadgar Pakdel, Mohammad Djaefar Moemenbellah-Fard\",\"doi\":\"10.1007/s10709-022-00164-8\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<p><p>The greenbottle blowfly Lucilia sericata (L. sericata) is increasingly used in larval therapy of chronic wounds. Netrins as bifunctional proteins are in the superfamily of Laminins secreted from larval salivary glands. The Netrin protein has a significant instructive role in axon guidance, causing neuronal outgrowth, angiogenesis, and cell migration. It seems to be crucial in wound healing and acts as a potential biomarker in diagnosing some clinical diseases. This survey aimed to identify molecular features and analyze in silico structural configuration of Netrin-A in L. sericata larvae. The larvae were reared under standard maggotarium conditions. The nucleic acid sequence of L. sericata Netrin-A (LSN-A) was then identified using rapid amplification of circular DNA ends (RACE) and rapid amplification of genomic ends (RAGE). Parts of the Netrin-A gene, including the middle, 3'-, and 5'-ends, were identified, TA cloned in pTG19 plasmid, and transferred into DH5ɑ Escherichia coli. Each part was sequenced and assembled using SeqMan software. This gene structure was further subjected to in silico analysis. The DNA of LSN-A was identified to be 2407 bp, while its mRNA sequence was recognized as 2115 bp by Oligo0.7 software. It translated the Netrin-A protein with 704 amino acid residues. Its estimated molecular weight was 78.6 kDa. Sequencing of this fragment and its BLAST analysis revealed laminin-based high (95%) similarity with the mRNA sequence of Lucilia cuprina Netrin-A. The 3-D structure of Netrin-A drawn by SWISS-MODEL exhibited its partial resemblance to the reference molecule Netrin-1 of Homo sapiens. This study supports the molecular and structural analyses of LSN-A protein, which could lead to wound treatment. Ultimately, it can be an effective candidate to ameliorate injury. Our next attempt is to produce LSN-A recombinant protein for use in biomedical sciences.</p>\",\"PeriodicalId\":1,\"journal\":{\"name\":\"Accounts of Chemical Research\",\"volume\":null,\"pages\":null},\"PeriodicalIF\":16.4000,\"publicationDate\":\"2022-12-01\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"\",\"citationCount\":\"0\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"Accounts of Chemical Research\",\"FirstCategoryId\":\"99\",\"ListUrlMain\":\"https://doi.org/10.1007/s10709-022-00164-8\",\"RegionNum\":1,\"RegionCategory\":\"化学\",\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"2022/9/22 0:00:00\",\"PubModel\":\"Epub\",\"JCR\":\"Q1\",\"JCRName\":\"CHEMISTRY, MULTIDISCIPLINARY\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"Accounts of Chemical Research","FirstCategoryId":"99","ListUrlMain":"https://doi.org/10.1007/s10709-022-00164-8","RegionNum":1,"RegionCategory":"化学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"2022/9/22 0:00:00","PubModel":"Epub","JCR":"Q1","JCRName":"CHEMISTRY, MULTIDISCIPLINARY","Score":null,"Total":0}
Identification, molecular characterization, and in silico structural analysis of larval salivary glands Netrin-A as a potent biomarker from Lucilia sericata (Diptera: Calliphoridae).
The greenbottle blowfly Lucilia sericata (L. sericata) is increasingly used in larval therapy of chronic wounds. Netrins as bifunctional proteins are in the superfamily of Laminins secreted from larval salivary glands. The Netrin protein has a significant instructive role in axon guidance, causing neuronal outgrowth, angiogenesis, and cell migration. It seems to be crucial in wound healing and acts as a potential biomarker in diagnosing some clinical diseases. This survey aimed to identify molecular features and analyze in silico structural configuration of Netrin-A in L. sericata larvae. The larvae were reared under standard maggotarium conditions. The nucleic acid sequence of L. sericata Netrin-A (LSN-A) was then identified using rapid amplification of circular DNA ends (RACE) and rapid amplification of genomic ends (RAGE). Parts of the Netrin-A gene, including the middle, 3'-, and 5'-ends, were identified, TA cloned in pTG19 plasmid, and transferred into DH5ɑ Escherichia coli. Each part was sequenced and assembled using SeqMan software. This gene structure was further subjected to in silico analysis. The DNA of LSN-A was identified to be 2407 bp, while its mRNA sequence was recognized as 2115 bp by Oligo0.7 software. It translated the Netrin-A protein with 704 amino acid residues. Its estimated molecular weight was 78.6 kDa. Sequencing of this fragment and its BLAST analysis revealed laminin-based high (95%) similarity with the mRNA sequence of Lucilia cuprina Netrin-A. The 3-D structure of Netrin-A drawn by SWISS-MODEL exhibited its partial resemblance to the reference molecule Netrin-1 of Homo sapiens. This study supports the molecular and structural analyses of LSN-A protein, which could lead to wound treatment. Ultimately, it can be an effective candidate to ameliorate injury. Our next attempt is to produce LSN-A recombinant protein for use in biomedical sciences.
期刊介绍：
Accounts of Chemical Research presents short, concise and critical articles offering easy-to-read overviews of basic research and applications in all areas of chemistry and biochemistry. These short reviews focus on research from the author’s own laboratory and are designed to teach the reader about a research project. In addition, Accounts of Chemical Research publishes commentaries that give an informed opinion on a current research problem. Special Issues online are devoted to a single topic of unusual activity and significance.
Accounts of Chemical Research replaces the traditional article abstract with an article "Conspectus." These entries synopsize the research affording the reader a closer look at the content and significance of an article. Through this provision of a more detailed description of the article contents, the Conspectus enhances the article's discoverability by search engines and the exposure for the research.
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