Expression and purification of the extracellular domains of human glycoprotein VI (GPVI) and the receptor for advanced glycation end products (RAGE) from Rattus norvegicus in Leishmania tarentolae

• Published in: Preparative biochemistry & biotechnology Vol. 47; no. 10; pp. 1008 - 1015
• Main Authors: Langer, Thomas, Corvey, Carsten, Kroll, Katja, Boscheinen, Oliver, Wendrich, Thomas, Dittrich, Werner
• Format: Journal Article
• Language: English
• Published: England Taylor & Francis 26.11.2017; Taylor & Francis Ltd
• Subjects: Advanced glycosylation end products; Animals; Bacteria; Biotechnology; Cloning, Molecular; Gene Expression; Glycoprotein VI; Glycoproteins; Glycosylation; GPVI; Humans; Insects; Leishmania - genetics; Leishmania tarentolae; Platelet Membrane Glycoproteins - chemistry; Platelet Membrane Glycoproteins - genetics; Platelet Membrane Glycoproteins - isolation & purification; Post-translation; Protein Domains; Protein folding; Proteins; Protozoa; RAGE; Rats; receptor for advanced glycation end products; Receptor for Advanced Glycation End Products - chemistry; Receptor for Advanced Glycation End Products - genetics; Receptor for Advanced Glycation End Products - isolation & purification; Recombinant Proteins - chemistry; Recombinant Proteins - genetics; Recombinant Proteins - isolation & purification; Translation; Yeast; Leishmania tarentolae; Glycoprotein VI; glycosylation; RAGE; receptor for advanced glycation end products; GPVI
• ISSN: 1082-6068; 1532-2297
• DOI: 10.1080/10826068.2017.1365252

Glycosylation is one of the most complex post-translational modifications and may have significant influence on the proper function of the corresponding proteins. Bacteria and yeast are, because of easy handling and cost reasons, the most frequently used systems for recombinant protein expression. Bacteria generally do not glycosylate proteins and yeast might tend to hyperglycosylate. Insect cell- and mammalian cell-based expression systems are able to produce complex N-glycosylation structures but are more complex to handle and more expensive. The nonpathogenic protozoa Leishmania tarentolae is an easy-to-handle alternative expression system for production of proteins requiring the eukaryotic protein folding machinery and post-translational modifications. We used and evaluated the system for the secretory expression of extracellular domains from human glycoprotein VI and the receptor for advanced glycation end products from rat. Both proteins were well expressed and homogeneously glycosylated. Analysis of the glycosylation pattern identified the structure as the conserved core pentasaccharide Man 3 GlcNac 2 .