Production and N-glycosylation of recombinant human cell adhesion molecule L1 from insect cells using the stable expression system. Effect of dimethyl sulfoxide

L1 is a cell adhesion molecule that is heavily glycosylated and is essential for normal development of the central nervous system. In this work, we compare the N-glycosylation of the L1 mutant that consists of immunoglobulin domains 5 and 6 (L1/Ig5-6), expressed in insect Spodoptera frugiperda Sf9 and Trichoplusia ni Tn cells, using the stable expression system. L1/Ig5-6 levels of 30 and 8 mg l(-1) were obtained from the two cell lines, respectively. The N-glycans were characterized by high-performance anion-exchange-chromatography with pulsed-amperometric-detection and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. The N-glycans from Sf9 cells were more homogeneous and consisted predominantly of the paucimannose-type structure Man alpha 6(Man alpha 3)Man beta 4GlcNAc beta 4(Fuc alpha 6)GlcNAc. On the other hand, the N-glycans from To cells were more heterogeneous and consisted of paucimannosetype structures with or without terminal N-acetylglucosamine. Allergenic Proximal alpha-linked fucose was only found in Tn cells. Dimethyl sulfoxide at 1.5% concentration has been found to increase the levels of L1/Ig5-6 and the L1 ectodomain in the Sf9 and To cells, without affecting cell viability nor protein integrity. Furthermore, the N-glycan composition of L1/lg5-6 was not affected by dimethyl sulfoxide, with only a slight increase in the percentage of the minor high-mannose-type structures. (C) 2009 Elsevier B.V. All rights reserved.