Purification and characterization of Alpha-Fetoprotein from the human hepatoblastoma HepG2 cell line in serum-free medium |
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Authors: | Patrizia Carlini Pasquale Ferranti Francesca Polizio Maria R Ciriolo Giuseppe Rotilio |
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Institution: | (1) Department of Biology, University of Rome “Tor Vergata”, Rome, Italy;(2) Department of Food Science, University of Naples “Federico II”, Portici, Italy |
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Abstract: | Alpha-fetoprotein (AFP) is a tumor-associated embryonic molecule whose precise biological function remains unclear. A complete
definition of the physiological activities of this oncofetal protein has been severely limited, until now, by the lack of
a purification procedure appropriate to obtain pure AFP in appreciable amount. The present report describes a purification
procedure extremely rapid and simple and takes advantage of the well-known fact that AFP contains copper. We have developed
a single-step purification procedure by immobilized copper-chelate affinity chromatography using the culture medium from human
hepatoblastoma cell line HepG2 grown in the absence of serum. This method yields AFP at high purity and high yield. Purified
AFP amino acid sequence, molecular mass, carbohydrate structure, and copper content were found to be in line with previous
studies. Moreover, we found that the purified AFP has superoxide dismutase activity with efficiency similar to that of the
native Cu, Zn SODs at physiological pH. This result may provide further support to the idea that AFP is a bifunctional protein,
acting in cellular defence against oxidative stress both as a copper buffer and as a superoxide radical scavenger. |
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Keywords: | Serum-free medium Human hepatoblastoma Alpha-fetoprotein Copper Superoxide dismutase |
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