title: Purification of the extracellular domain of the membrane protein GlialCAM expressed in HEK and CHO cells and comparison of the glycosylation.
abstract: Adhesion molecules are essential for a wide range of biological and physiological functions, including cell-cell interactions, cell interactions with the extracellular matrix, cell migration, proliferation and survival. Defects in cell adhesion have been associated with pathological conditions such as neoplasia, and neurodegenerative diseases. We have identified a new adhesion molecule of the immunoglobulin family, GlialCAM. The same protein was recently published under the name hepaCAM and was suggested to be associated with hepatocellular carcinoma. Here we have expressed and purified the extracellular domain of this molecule in two mammalian expression systems, HEK and CHO cells. A three step purification protocol gave an over 95% pure protein. The extracellular domain of GlialCAM possesses several potential N- and O-glycosylation sites. Glycosylation is one of the most common post-translational modifications of secreted proteins and of the extracellular domains of membrane bound proteins. It can influence both the activity and the stability of the protein. The glycosylation pattern has been shown to depend on the cell type where the protein is expressed. We examined if differences in the glycosylation of this protein could be detected when it was expressed in the two commonly used mammalian expression systems, HEK and CHO. Differences in the glycosylation were detected.
authors: Geneva Research Centre, Merck Serono International S.A., 9 Chemin des Mines, 1202 Geneva, Switzerland.
authors: Jean-Philippe
authors: JP
authors: Gaudry
authors: Christian
authors: C
authors: Arod
authors: Christophe
authors: C
authors: Sauvage
authors: Stephane
authors: S
authors: Busso
authors: Philippe
authors: P
authors: Dupraz
authors: Renata
authors: R
authors: Pankiewicz
authors: Bruno
authors: B
authors: Antonsson
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D016466:CHO Cells; D015815:Cell Adhesion Molecules / Q000096:biosynthesis / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification; D018797:Cell Cycle Proteins; D002460:Cell Line; D006224:Cricetinae; D003412:Cricetulus; D006031:Glycosylation; D006801:Humans; D008969:Molecular Sequence Data; D011499:Protein Processing, Post-Translational; D017434:Protein Structure, Tertiary; D011506:Proteins / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000302:isolation & purification; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
chemical_list: D015815:Cell Adhesion Molecules; D018797:Cell Cycle Proteins; C487620:HEPACAM protein, human; D011506:Proteins; D011994:Recombinant Proteins