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Experimental study on the purification of human leptin / 中国组织工程研究
Chinese Journal of Tissue Engineering Research ; (53): 162-164, 2005.
Artículo en Chino | WPRIM | ID: wpr-409774
ABSTRACT

BACKGROUND:

Leptin is a hormone produced predominantly by adipocytes and has a variety of physiological functions. It has been a hot spot in energy metabolism research. However, leptin presently used is usually produced by E coli, in which leptin cDNA is expressed and is in the form of insoluble inclusion body. Therefore, extremely complicated reactivating procedure is needed to obtain biologically activated human leptin.

OBJECTIVE:

To explore the condition for purification of leptin in non-affinity chromatography in order to obtain soluble human leptin.

DESIGN:

An observational experiment.

SETTING:

Molecular laboratory of biochemical department in a medical college.MATERIALS The strong anion exchanger sepharose Q and hydrophobic phenyl sepharose 6 were used in different conditions for removal of as many contaminants as possible.

METHODS:

The supernatant of pichia pastoris yeast culture solution was first purified through Q column chromatography, the protein was collected and was further purified through hydrophobic support with 1 mol/ L (NH4) 2SO4.MAIN OUTCOME

MEASURES:

Gel scanning revealed that the purity of human leptin was 42. 3% before purification, 89.6% after Q column chromatography and 96.2% after hydrophobic interaction chromatography.

RESULTS:

The post-purification product presented a s. ingle band in SDS-PAGE. Gel scanning revealed that the purity of human leptin was 42.3% before purification, 89.6% after Q column chromatography and 96.2% after hydrophobic interaction chromatography.

CONCLUSION:

The combined use of strong anion exchange chromatography and hydrophobic interaction chromatography can effectively purify leptin expressed by pichia pastoris yeast and the purity is identical to that of nickel affinity column chromatography. It provides reliable evidence and method for possible manufacture of human leptin and lays experimental basis for leptin-related research.
Texto completo: Disponible Índice: WPRIM (Pacífico Occidental) Idioma: Chino Revista: Chinese Journal of Tissue Engineering Research Año: 2005 Tipo del documento: Artículo

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Texto completo: Disponible Índice: WPRIM (Pacífico Occidental) Idioma: Chino Revista: Chinese Journal of Tissue Engineering Research Año: 2005 Tipo del documento: Artículo