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Protein Expr Purif ; 58(1): 70-7, 2008 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-18068377

RESUMO

MGF is a product of a unique muscle-specific splice variant of IGF1 gene (insulin-like growth factor). Its peculiar feature is a specific E-peptide, a 16 a.a. strand at the C-terminus. MGF increases cellular proliferation and inhibits terminal differentiation of myoblasts necessary for the secondary myotube formation. Previous analysis of physiological effects of MGF was performed using indirect methods such as RT-PCR based examination of the transcript contents in normal tissues, adenovirus-mediated DNA delivery and synthetic E-domain administration. Here, we describe isolation and purification of recombinant MGF thus allowing for the first time the possibility of direct examining MGF effects. The recombinant MGF of directly examining--was expressed in Escherichia coli as inclusion bodies (about 100-200mg/l), purified and refolded. Biological activity of refolded MGF was analyzed in vitro in proliferation assays with normal human myoblasts. As a result of our work, it has become possible to generate a standard MGF control with characterized activity and a ready-to use MGF test-system neither of which have been previously described. Our data open opportunities for the future works on MGF characterization and to the development of a powerful and highly specific therapeutic agent potentially applicable for muscle growth up-regulation, post-trauma muscle repair, age and hereditary myodystrophy mitigation and in sport medicine.


Assuntos
Escherichia coli/genética , Fator de Transcrição STAT5/biossíntese , Proteínas Supressoras de Tumor/biossíntese , Sequência de Aminoácidos , Sequência de Bases , Proliferação de Células , Escherichia coli/metabolismo , Expressão Gênica , Humanos , Dados de Sequência Molecular , Mioblastos/citologia , Plasmídeos , Dobramento de Proteína , Proteínas Recombinantes/biossíntese , Proteínas Recombinantes/genética , Proteínas Recombinantes/isolamento & purificação , Fator de Transcrição STAT5/química , Fator de Transcrição STAT5/genética , Fator de Transcrição STAT5/farmacologia , Proteínas Supressoras de Tumor/química , Proteínas Supressoras de Tumor/genética , Proteínas Supressoras de Tumor/farmacologia
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