Protocol for Increasing the Sensitivity of MS-Based Protein Detection in Human Chorionic Villi
An important step in the proteomic analysis of missing proteins is the use of a wide range of tissues, optimal extraction, and the processing of protein material in order to ensure the highest sensitivity in downstream protein detection. This work describes a purification protocol for identifying lo...
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MDPI AG
2022-05-01
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| Series: | Current Issues in Molecular Biology |
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| Online Access: | https://www.mdpi.com/1467-3045/44/5/140 |
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| author | Timur Shkrigunov Pavel Pogodin Victor Zgoda Olesya Larina Yulia Kisrieva Maria Klimenko Oleg Latyshkevich Peter Klimenko Andrey Lisitsa Natalia Petushkova |
| author_facet | Timur Shkrigunov Pavel Pogodin Victor Zgoda Olesya Larina Yulia Kisrieva Maria Klimenko Oleg Latyshkevich Peter Klimenko Andrey Lisitsa Natalia Petushkova |
| author_sort | Timur Shkrigunov |
| collection | DOAJ |
| description | An important step in the proteomic analysis of missing proteins is the use of a wide range of tissues, optimal extraction, and the processing of protein material in order to ensure the highest sensitivity in downstream protein detection. This work describes a purification protocol for identifying low-abundance proteins in human chorionic villi using the proposed “1DE-gel concentration” method. This involves the removal of SDS in a short electrophoresis run in a stacking gel without protein separation. Following the in-gel digestion of the obtained holistic single protein band, we used the peptide mixture for further LC–MS/MS analysis. Statistically significant results were derived from six datasets, containing three treatments, each from two tissue sources (elective or missed abortions). The 1DE-gel concentration increased the coverage of the chorionic villus proteome. Our approach allowed the identification of 15 low-abundance proteins, of which some had not been previously detected via the mass spectrometry of trophoblasts. In the post hoc data analysis, we found a dubious or uncertain protein (PSG7) encoded on human chromosome 19 according to neXtProt. A proteomic sample preparation workflow with the 1DE-gel concentration can be used as a prospective tool for uncovering the low-abundance part of the human proteome. |
| first_indexed | 2024-03-10T03:05:35Z |
| format | Article |
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| institution | Directory Open Access Journal |
| issn | 1467-3037 1467-3045 |
| language | English |
| last_indexed | 2024-03-10T03:05:35Z |
| publishDate | 2022-05-01 |
| publisher | MDPI AG |
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| series | Current Issues in Molecular Biology |
| spelling | doaj.art-3b563a346b864c9ca9e73a4ce7b5c9e02023-11-23T10:32:18ZengMDPI AGCurrent Issues in Molecular Biology1467-30371467-30452022-05-014452069208810.3390/cimb44050140Protocol for Increasing the Sensitivity of MS-Based Protein Detection in Human Chorionic VilliTimur Shkrigunov0Pavel Pogodin1Victor Zgoda2Olesya Larina3Yulia Kisrieva4Maria Klimenko5Oleg Latyshkevich6Peter Klimenko7Andrey Lisitsa8Natalia Petushkova9Center of Scientific and Practical Education, Institute of Biomedical Chemistry, 119121 Moscow, RussiaLaboratory of Structure-Function Based Drug Design, Institute of Biomedical Chemistry, 119121 Moscow, RussiaLaboratory of Systems Biology, Institute of Biomedical Chemistry, 119121 Moscow, RussiaLaboratory of Microsomal Oxidation, Institute of Biomedical Chemistry, 119121 Moscow, RussiaLaboratory of Microsomal Oxidation, Institute of Biomedical Chemistry, 119121 Moscow, RussiaCenter for Family Planning and Reproduction, Moscow Department of Health, 117209 Moscow, RussiaCenter for Family Planning and Reproduction, Moscow Department of Health, 117209 Moscow, RussiaDepartment of Obstetrics and Gynecology, Pirogov Russian National Research Medical University, 117997 Moscow, RussiaCenter of Scientific and Practical Education, Institute of Biomedical Chemistry, 119121 Moscow, RussiaLaboratory of Microsomal Oxidation, Institute of Biomedical Chemistry, 119121 Moscow, RussiaAn important step in the proteomic analysis of missing proteins is the use of a wide range of tissues, optimal extraction, and the processing of protein material in order to ensure the highest sensitivity in downstream protein detection. This work describes a purification protocol for identifying low-abundance proteins in human chorionic villi using the proposed “1DE-gel concentration” method. This involves the removal of SDS in a short electrophoresis run in a stacking gel without protein separation. Following the in-gel digestion of the obtained holistic single protein band, we used the peptide mixture for further LC–MS/MS analysis. Statistically significant results were derived from six datasets, containing three treatments, each from two tissue sources (elective or missed abortions). The 1DE-gel concentration increased the coverage of the chorionic villus proteome. Our approach allowed the identification of 15 low-abundance proteins, of which some had not been previously detected via the mass spectrometry of trophoblasts. In the post hoc data analysis, we found a dubious or uncertain protein (PSG7) encoded on human chromosome 19 according to neXtProt. A proteomic sample preparation workflow with the 1DE-gel concentration can be used as a prospective tool for uncovering the low-abundance part of the human proteome.https://www.mdpi.com/1467-3045/44/5/140chorionic villiSDS extracts1DE-gel concentrationLC–MS/MSlow-abundance proteinselective abortion |
| spellingShingle | Timur Shkrigunov Pavel Pogodin Victor Zgoda Olesya Larina Yulia Kisrieva Maria Klimenko Oleg Latyshkevich Peter Klimenko Andrey Lisitsa Natalia Petushkova Protocol for Increasing the Sensitivity of MS-Based Protein Detection in Human Chorionic Villi Current Issues in Molecular Biology chorionic villi SDS extracts 1DE-gel concentration LC–MS/MS low-abundance proteins elective abortion |
| title | Protocol for Increasing the Sensitivity of MS-Based Protein Detection in Human Chorionic Villi |
| title_full | Protocol for Increasing the Sensitivity of MS-Based Protein Detection in Human Chorionic Villi |
| title_fullStr | Protocol for Increasing the Sensitivity of MS-Based Protein Detection in Human Chorionic Villi |
| title_full_unstemmed | Protocol for Increasing the Sensitivity of MS-Based Protein Detection in Human Chorionic Villi |
| title_short | Protocol for Increasing the Sensitivity of MS-Based Protein Detection in Human Chorionic Villi |
| title_sort | protocol for increasing the sensitivity of ms based protein detection in human chorionic villi |
| topic | chorionic villi SDS extracts 1DE-gel concentration LC–MS/MS low-abundance proteins elective abortion |
| url | https://www.mdpi.com/1467-3045/44/5/140 |
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