Expression and purification of SARS coronavirus proteins using SUMO-fusions.
Protein Expr Purif
; 42(1): 100-10, 2005 Jul.
Article
em En
| MEDLINE
| ID: mdl-15939295
ABSTRACT
Severe acute respiratory syndrome coronavirus (SARS-CoV) proteins belong to a large group of proteins that is difficult to express in traditional expression systems. The ability to express and purify SARS-CoV proteins in large quantities is critical for basic research and for development of pharmaceutical agents. The work reported here demonstrates (1) fusion of SUMO (small ubiquitin-related modifier), a 100 amino acid polypeptide, to the N-termini of SARS-CoV proteins dramatically enhances expression in Escherichia coli cells and (2) 6x His-tagged SUMO-fusions facilitate rapid purification of the viral proteins on a large scale. We have exploited the natural chaperoning properties of SUMO to develop an expression system suitable for proteins that cannot be expressed by traditional methodologies. A unique feature of the system is the SUMO tag, which enhances expression, facilitates purification, and can be efficiently cleaved by a SUMO-specific protease to generate native protein with a desired N-terminus. We have purified various SARS-CoV proteins under either native or denaturing conditions. These purified proteins have been used to generate highly specific polyclonal antibodies. Our study suggests that the SUMO-fusion technology will be useful for enhancing expression and purification of the viral proteins for structural and functional studies as well as for therapeutic uses.
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Proteínas Virais
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Proteínas Recombinantes de Fusão
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Expressão Gênica
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Proteínas Modificadoras Pequenas Relacionadas à Ubiquitina
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Coronavírus Relacionado à Síndrome Respiratória Aguda Grave
Idioma:
En
Ano de publicação:
2005
Tipo de documento:
Article