In a previous paper, the biological activity of a 216-amino acid recombinant truncated form of the soybean 7S globulin alfa' subunit, known to control cholesterol and triglyceride homeostasis, was described. In this work, a shorter version of the polypeptide chain, spanning 142 amino acid residues from the N-terminus and thus exclusively including the so-called extension region, was cloned and over-expressed in Pichia pastoris. The yield of the recombinant polypeptide, which was termed alfa'E, was 8-fold greater than the previous truncated version. The alfa'E polypeptide was puriﬁed by simple conventional biochemical techniques to make it available for biological assays. Human hepatoma cell lines (Hep G2) were used to monitor the uptake and degradation of labeled low-density lipoproteins (LDL), according to an established procedure. The LDL uptake (+86%) and degradation (+94%) by cells tested at the highest alfa'E dose (2 microM) were similar to those found in cells incubated with 1 microM simvastatin, a potent inhibitor of cholesterol biosynthesis. Additionally, the cell response to alfa'E was found to be dose-dependent. The present ﬁndings strongly suggest that this recombinant polypeptide, or a fragment thereof, is the molecular determinant for cholesterol homeostasis and open new prospects for understanding the mechanism involved in this biological response, as a gateway to its utilization in lipid-lowering therapies.
|Titolo:||Heterologous expression and purification of the soybean 7S globulin α′ subunit extension region: In vitro evidence of its involvement in cell cholesterol homeostasis|
|Parole Chiave:||7S soybean globulin ; alfa' chain ; extension region ; Hep G2 cells ; LDL-receptors ; cell cholesterol homeostasis|
|Settore Scientifico Disciplinare:||Settore BIO/14 - Farmacologia|
Settore BIO/10 - Biochimica
|Data di pubblicazione:||2011|
|Digital Object Identifier (DOI):||http://dx.doi.org/10.1016/j.pep.2011.07.008|
|Appare nelle tipologie:||01 - Articolo su periodico|