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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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Autores principales: Shkrigunov, Timur, Pogodin, Pavel, Zgoda, Victor, Larina, Olesya, Kisrieva, Yulia, Klimenko, Maria, Latyshkevich, Oleg, Klimenko, Peter, Lisitsa, Andrey, Petushkova, Natalia
Formato: Online Artículo Texto
Lenguaje:English
Publicado: MDPI 2022
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9164042/
https://www.ncbi.nlm.nih.gov/pubmed/35678669
http://dx.doi.org/10.3390/cimb44050140
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author Shkrigunov, Timur
Pogodin, Pavel
Zgoda, Victor
Larina, Olesya
Kisrieva, Yulia
Klimenko, Maria
Latyshkevich, Oleg
Klimenko, Peter
Lisitsa, Andrey
Petushkova, Natalia
author_facet Shkrigunov, Timur
Pogodin, Pavel
Zgoda, Victor
Larina, Olesya
Kisrieva, Yulia
Klimenko, Maria
Latyshkevich, Oleg
Klimenko, Peter
Lisitsa, Andrey
Petushkova, Natalia
author_sort Shkrigunov, Timur
collection PubMed
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.
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spelling pubmed-91640422022-06-04 Protocol for Increasing the Sensitivity of MS-Based Protein Detection in Human Chorionic Villi Shkrigunov, Timur Pogodin, Pavel Zgoda, Victor Larina, Olesya Kisrieva, Yulia Klimenko, Maria Latyshkevich, Oleg Klimenko, Peter Lisitsa, Andrey Petushkova, Natalia Curr Issues Mol Biol Article 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. MDPI 2022-05-09 /pmc/articles/PMC9164042/ /pubmed/35678669 http://dx.doi.org/10.3390/cimb44050140 Text en © 2022 by the authors. https://creativecommons.org/licenses/by/4.0/Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/).
spellingShingle Article
Shkrigunov, Timur
Pogodin, Pavel
Zgoda, Victor
Larina, Olesya
Kisrieva, Yulia
Klimenko, Maria
Latyshkevich, Oleg
Klimenko, Peter
Lisitsa, Andrey
Petushkova, Natalia
Protocol for Increasing the Sensitivity of MS-Based Protein Detection in Human Chorionic Villi
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 Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9164042/
https://www.ncbi.nlm.nih.gov/pubmed/35678669
http://dx.doi.org/10.3390/cimb44050140
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