Divide-and-Conquer Matrisome Protein (DC-MaP) Strategy: An MS-Friendly Approach to Proteomic Matrisome Characterization.
Int J Mol Sci
; 21(23)2020 11 30.
Article
em En
| MEDLINE
| ID: mdl-33266304
ABSTRACT
Currently, the extracellular matrix (ECM) is considered a pivotal complex meshwork of macromolecules playing a plethora of biomolecular functions in health and disease beyond its commonly known mechanical role. Only by unraveling its composition can we leverage related tissue engineering and pharmacological efforts. Nevertheless, its unbiased proteomic identification still encounters some limitations mainly due to partial ECM enrichment by precipitation, sequential fractionation using unfriendly-mass spectrometry (MS) detergents, and resuspension with harsh reagents that need to be entirely removed prior to analysis. These methods can be technically challenging and labor-intensive, which affects the reproducibility of ECM identification and induces protein loss. Here, we present a simple new method applicable to tissue fragments of 10 mg and more. The technique has been validated on human ovarian tissue and involves a standardized procedure for sample processing with an MS-compatible detergent and combined centrifugation. This two-step protocol eliminates the need for laborious sample clarification and divides our samples into 2 fractions, soluble and insoluble, successively enriched with matrisome-associated (ECM-interacting) and core matrisome (structural ECM) proteins.
Palavras-chave
Texto completo:
1
Base de dados:
MEDLINE
Assunto principal:
Proteínas da Matriz Extracelular
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Proteoma
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Proteômica
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Matriz Extracelular
Tipo de estudo:
Prognostic_studies
Limite:
Humans
Idioma:
En
Ano de publicação:
2020
Tipo de documento:
Article