Abstract
Purifying progranulin may be useful in a variety of situations, for example, after it has been mutated or otherwise modified or when working with a species for which commercially produced progranulin is unavailable. A method to express and purify human progranulin is presented. Progranulin is transiently expressed in mammalian cells and isolated from their conditioned medium before purification by reversed-phase high-performance liquid chromatography (RP-HPLC). Human progranulin is used as an example, but the protocol can be applied to any other progranulin protein. Modifications of the expression-purification strategy for metabolic labeling of progranulin and analytical systems based on heparin-affinity chromatography are presented.
This is a preview of subscription content, log in via an institution to check access.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
Cenik B, Sephton CF, Cenik BK et al (2012) Progranulin: a proteolytically processed protein at the crossroads of inflammation and neurodegeneration. J Biol Chem 287:32298–32306
Hu F, Padukkavidana T, Vægter CB, Brady OA, Zheng Y, Mackenzie IR, Feldman HH, Nykjaer A, Strittmatter SM (2010) Sortilin-mediated endocytosis determines levels of the frontotemporal dementia protein, progranulin. Neuron 68:654–667
Tanimoto R, Morcavallo A, Terracciano M, Xu SQ, Stefanello M, Buraschi S, Lu KG, Bagley DH, Gomella LG, Scotlandi K, Belfiore A, Iozzo RV, Morrione A (2015) Sortilin regulates progranulin action in castration-resistant prostate cancer cells. Endocrinology 156:58–70
Zheng Y, Brady OA, Meng PS, Mao Y, Hu F (2011) C-terminus of progranulin interacts with the beta-propeller region of sortilin to regulate progranulin trafficking. PLoS One 6:e21023
He Z, Bateman A (1999) Progranulin gene expression regulates epithelial cell growth and promotes tumor growth in vivo. Cancer Res 59:3222–3229
He Z, Ismail A, Kriazhev L, Sadvakassova G et al (2002) Progranulin (PC-cell-derived growth factor/acrogranin) regulates invasion and cell survival. Cancer Res 62:5590–5596
Gluzman Y (1981) SV40-transformed simian cells support the replication of early SV40 mutants. Cell 23:175–182
Ohki EC, Tilkins ML, Ciccarone VC et al (2001) Improving the transfection efficiency of post-mitotic neurons. J Neurosci Methods 112:95–99
Dalby B, Cates S, Harris A et al (2004) Advanced transfection with Lipofectamine 2000 reagent: primary neurons, siRNA, and high-throughput applications. Methods 33:95–103
James S, Gibbs BF, Toney K, Bennett HP (1994) Purification of antimicrobial peptides from an extract of the skin of Xenopus laevis using heparin-affinity HPLC: characterization by ion-spray mass spectrometry. Anal Biochem 217:84–90
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2018 Springer Science+Business Media, LLC, part of Springer Nature
About this protocol
Cite this protocol
Zhao, C.Y., Bateman, A. (2018). Methods for Expression and Purification of Biologically Active Recombinant Progranulin. In: Bateman, A., Bennett, H., Cheung, S. (eds) Progranulin. Methods in Molecular Biology, vol 1806. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-8559-3_3
Download citation
DOI: https://doi.org/10.1007/978-1-4939-8559-3_3
Published:
Publisher Name: Humana Press, New York, NY
Print ISBN: 978-1-4939-8557-9
Online ISBN: 978-1-4939-8559-3
eBook Packages: Springer Protocols