BacMam production and crystal structure of nonglycosylated apo human furin at 1.89â
Å resolution.
Acta Crystallogr F Struct Biol Commun
; 75(Pt 4): 239-245, 2019 Apr 01.
Article
en En
| MEDLINE
| ID: mdl-30950824
ABSTRACT
Furin, also called proprotein convertase subtilisin/kexin 3 (PCSK3), is a calcium-dependent serine endoprotease that processes a wide variety of proproteins involved in cell function and homeostasis. Dysregulation of furin has been implicated in numerous disease states, including cancer and fibrosis. Mammalian cell expression of the furin ectodomain typically produces a highly glycosylated, heterogeneous protein, which can make crystallographic studies difficult. Here, the expression and purification of nonglycosylated human furin using the BacMam technology and site-directed mutagenesis of the glycosylation sites is reported. Nonglycosylated furin produced using this system retains full proteolytic activity indistinguishable from that of the glycosylated protein. Importantly, the nonglycosylated furin protein reliably forms extremely durable apo crystals that diffract to high resolution. These crystals can be soaked with a wide variety of inhibitors to enable a structure-guided drug-discovery campaign.
Palabras clave
Texto completo:
1
Banco de datos:
MEDLINE
Asunto principal:
Apoproteínas
/
Bioquímica
/
Furina
Límite:
Animals
/
Humans
Idioma:
En
Revista:
Acta Crystallogr F Struct Biol Commun
Año:
2019
Tipo del documento:
Article
País de afiliación:
Estados Unidos