L. Pagani, C. Chinello, A. Mahajneh, F. Clerici, Lucrezia Criscuolo, A. Favalli, P. Gruarin, R. Grifantini, A. Bandera, A. Lombardi, R. Ungaro, A. Muscatello, F. Blasi, A. Gori, F. Magni
{"title":"非靶向质谱法研究人血浆和唾液蛋白质组中SARS-CoV-2蛋白","authors":"L. Pagani, C. Chinello, A. Mahajneh, F. Clerici, Lucrezia Criscuolo, A. Favalli, P. Gruarin, R. Grifantini, A. Bandera, A. Lombardi, R. Ungaro, A. Muscatello, F. Blasi, A. Gori, F. Magni","doi":"10.3390/biochem2010005","DOIUrl":null,"url":null,"abstract":"Since the start of the COVID-19 outbreak, more than four million people have died of this disease. Given its ability to provide a precise response, mass spectrometry-based proteomics could represent a useful tool to study this pathology. To this end, an untargeted nLC-ESI-MS/MS-based method to characterise SARS-CoV-2 proteins, including possible variants, and investigate human saliva and plasma proteome in a single analysis was developed for further application in patients. Four SARS-CoV-2 recombinant proteins, three (S1–S2–RBD) belonging to the spike glycoprotein (S) and one corresponding to the nucleoprotein (N), were prepared and analysed with nLC-UHRTOF by injecting decreasing amounts to establish the limit of detection (LOD) of the method. This was determined as 10 pg for all the components of the S protein and for N (71 amol and 213 amol, respectively). Various viral inactivation strategies plus deglycosylation and digestion approaches were then tested in saliva and plasma spiked with different quantities of SARS-CoV-2 recombinant proteins. The limit of characterisation (LOC) in saliva for the N and S proteins was observed at 100 pg (coverage of 20% and 3%, respectively); instead, in plasma, it was 33 pg for N and 330 pg for the S protein, with a coverage of 4% for both. About 300 and 800 human proteins were identified in plasma and saliva, respectively, including several key effectors and pathways that are known to be altered in COVID-19 patients. In conclusion, this approach allows SARS-CoV-2 proteins and the human proteome to be simultaneously explored, both for plasma and saliva, showing a high relevant potential for retrospective studies aimed at investigating possible virus variants and for patient stratification.","PeriodicalId":72357,"journal":{"name":"BioChem","volume":null,"pages":null},"PeriodicalIF":0.0000,"publicationDate":"2022-02-08","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"2","resultStr":"{\"title\":\"Untargeted Mass Spectrometry Approach to Study SARS-CoV-2 Proteins in Human Plasma and Saliva Proteome\",\"authors\":\"L. Pagani, C. Chinello, A. Mahajneh, F. Clerici, Lucrezia Criscuolo, A. Favalli, P. Gruarin, R. Grifantini, A. Bandera, A. Lombardi, R. Ungaro, A. Muscatello, F. Blasi, A. Gori, F. Magni\",\"doi\":\"10.3390/biochem2010005\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"Since the start of the COVID-19 outbreak, more than four million people have died of this disease. Given its ability to provide a precise response, mass spectrometry-based proteomics could represent a useful tool to study this pathology. To this end, an untargeted nLC-ESI-MS/MS-based method to characterise SARS-CoV-2 proteins, including possible variants, and investigate human saliva and plasma proteome in a single analysis was developed for further application in patients. Four SARS-CoV-2 recombinant proteins, three (S1–S2–RBD) belonging to the spike glycoprotein (S) and one corresponding to the nucleoprotein (N), were prepared and analysed with nLC-UHRTOF by injecting decreasing amounts to establish the limit of detection (LOD) of the method. This was determined as 10 pg for all the components of the S protein and for N (71 amol and 213 amol, respectively). Various viral inactivation strategies plus deglycosylation and digestion approaches were then tested in saliva and plasma spiked with different quantities of SARS-CoV-2 recombinant proteins. The limit of characterisation (LOC) in saliva for the N and S proteins was observed at 100 pg (coverage of 20% and 3%, respectively); instead, in plasma, it was 33 pg for N and 330 pg for the S protein, with a coverage of 4% for both. About 300 and 800 human proteins were identified in plasma and saliva, respectively, including several key effectors and pathways that are known to be altered in COVID-19 patients. In conclusion, this approach allows SARS-CoV-2 proteins and the human proteome to be simultaneously explored, both for plasma and saliva, showing a high relevant potential for retrospective studies aimed at investigating possible virus variants and for patient stratification.\",\"PeriodicalId\":72357,\"journal\":{\"name\":\"BioChem\",\"volume\":null,\"pages\":null},\"PeriodicalIF\":0.0000,\"publicationDate\":\"2022-02-08\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"\",\"citationCount\":\"2\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"BioChem\",\"FirstCategoryId\":\"1085\",\"ListUrlMain\":\"https://doi.org/10.3390/biochem2010005\",\"RegionNum\":0,\"RegionCategory\":null,\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"\",\"PubModel\":\"\",\"JCR\":\"\",\"JCRName\":\"\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"BioChem","FirstCategoryId":"1085","ListUrlMain":"https://doi.org/10.3390/biochem2010005","RegionNum":0,"RegionCategory":null,"ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"","JCRName":"","Score":null,"Total":0}
Untargeted Mass Spectrometry Approach to Study SARS-CoV-2 Proteins in Human Plasma and Saliva Proteome
Since the start of the COVID-19 outbreak, more than four million people have died of this disease. Given its ability to provide a precise response, mass spectrometry-based proteomics could represent a useful tool to study this pathology. To this end, an untargeted nLC-ESI-MS/MS-based method to characterise SARS-CoV-2 proteins, including possible variants, and investigate human saliva and plasma proteome in a single analysis was developed for further application in patients. Four SARS-CoV-2 recombinant proteins, three (S1–S2–RBD) belonging to the spike glycoprotein (S) and one corresponding to the nucleoprotein (N), were prepared and analysed with nLC-UHRTOF by injecting decreasing amounts to establish the limit of detection (LOD) of the method. This was determined as 10 pg for all the components of the S protein and for N (71 amol and 213 amol, respectively). Various viral inactivation strategies plus deglycosylation and digestion approaches were then tested in saliva and plasma spiked with different quantities of SARS-CoV-2 recombinant proteins. The limit of characterisation (LOC) in saliva for the N and S proteins was observed at 100 pg (coverage of 20% and 3%, respectively); instead, in plasma, it was 33 pg for N and 330 pg for the S protein, with a coverage of 4% for both. About 300 and 800 human proteins were identified in plasma and saliva, respectively, including several key effectors and pathways that are known to be altered in COVID-19 patients. In conclusion, this approach allows SARS-CoV-2 proteins and the human proteome to be simultaneously explored, both for plasma and saliva, showing a high relevant potential for retrospective studies aimed at investigating possible virus variants and for patient stratification.