Microfluidic protein isolation and sample preparation for high-resolution cryo-EM.
Proc Natl Acad Sci U S A
; 116(30): 15007-15012, 2019 07 23.
Article
em En
| MEDLINE
| ID: mdl-31292253
ABSTRACT
High-resolution structural information is essential to understand protein function. Protein-structure determination needs a considerable amount of protein, which can be challenging to produce, often involving harsh and lengthy procedures. In contrast, the several thousand to a few million protein particles required for structure determination by cryogenic electron microscopy (cryo-EM) can be provided by miniaturized systems. Here, we present a microfluidic method for the rapid isolation of a target protein and its direct preparation for cryo-EM. Less than 1 µL of cell lysate is required as starting material to solve the atomic structure of the untagged, endogenous human 20S proteasome. Our work paves the way for high-throughput structure determination of proteins from minimal amounts of cell lysate and opens more opportunities for the isolation of sensitive, endogenous protein complexes.
Palavras-chave
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Processamento de Imagem Assistida por Computador
/
Microscopia Crioeletrônica
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Subunidades Proteicas
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Complexo de Endopeptidases do Proteassoma
Limite:
Humans
Idioma:
En
Revista:
Proc Natl Acad Sci U S A
Ano de publicação:
2019
Tipo de documento:
Article
País de afiliação:
Suíça