Recombinant human pigment epithelium-derived factor (PEDF): characterization of PEDF overexpressed and secreted by eukaryotic cells

Protein Science : a Publication of the Protein Society
E StratikosS Patricia Becerra

Abstract

Pigment epithelium-derived factor (PEDF) is a serpin found in the interphotoreceptor matrix of the eye, which, although not a proteinase inhibitor, possesses a number of important biological properties, including promotion of neurite outgrowth and differential expression in quiescent versus senescent states of certain cell types. The low amounts present in the eye, together with the impracticality of using the eye as a source for isolation of the human protein, make it important to establish a system for overexpression of the recombinant protein for biochemical and biological studies. We describe here the expression and secretion of full-length glycosylated human recombinant PEDF at high levels (> 20 micrograms/ mL) into the growth medium of baby hamster kidney cells and characterization of the purified rPEDF by circular dichroism and fluorescence spectroscopies and neurite outgrowth assay. By these assays, the recombinant protein behaves as expected for a correctly folded full-length human PEDF. The availability of milligram amounts of PEDF has permitted quantitation of its heparin binding properties and of the effect of reactive center cleavage on the stability of PEDF towards thermal and guanidine hydrochloride denaturation.

References

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Citations

Aug 5, 1998·Biochimica Et Biophysica Acta·L A Perez-MediavillaS P Becerra
Mar 15, 2013·Nature Reviews. Cancer·S Patricia Becerra, Vicente Notario
Sep 23, 2008·The Journal of Biological Chemistry·S Patricia BecerraJoe G Hollyfield
Jun 16, 2012·Journal of Biomedicine & Biotechnology·P SubramanianS P Becerra
Apr 17, 2012·International Journal of Oncology·Monika DeshpandeS Patricia Becerra
Apr 21, 2004·Proceedings of the National Academy of Sciences of the United States of America·Hua LiuPatrick Y Tong
Sep 20, 2001·Proceedings of the National Academy of Sciences of the United States of America·M SimonovicK Volz
Feb 20, 2010·Growth Factors·Matthew L BroadheadCrispin R Dass
Jan 10, 2016·Journal of Experimental & Clinical Cancer Research : CR·Louiza Belkacemi, Shaun Xiaoliu Zhang
May 14, 2004·The Journal of Investigative Dermatology·Mary Kay FrancisVincent J Cristofalo
Oct 17, 2009·Cancer Science·Huasheng YangGuoquan Gao
Apr 24, 2010·The FEBS Journal·Luigi NotariS P Becerra
Jun 14, 2002·Biochemical and Biophysical Research Communications·Helen HutchingsJean Plouët
Jul 15, 2015·American Journal of Physiology. Heart and Circulatory Physiology·Mateusz S WietechaLuisa A DiPietro
Nov 26, 1998·International Journal of Developmental Neuroscience : the Official Journal of the International Society for Developmental Neuroscience·F Malchiodi-AlbediJ Tombran-Tink
Sep 19, 2002·The Journal of Biological Chemistry·Christina MeyerS Patricia Becerra
Jan 5, 2007·The Journal of Biological Chemistry·Zuzana ValnickovaJan J Enghild
Jul 26, 2018·Scientific Reports·Elizabeth R MichalczykLuisa A DiPietro
Oct 4, 2020·International Journal of Molecular Sciences·Francisco J Valiente-SorianoManuel Vidal-Sanz
Dec 12, 2002·Chemical Reviews·Peter G W Gettins

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