beta-Secretase (PSEC) was expressed in Trichoplusia ni BTI Tn5B1-4(Tn5B1-4) cells transformed with cDNAs encoding beta 1,4-galactosyl-transferase (GaIT) and Gal beta 1,4-GlcNAc alpha 2,6-sialyltransferase (ST). The apparent molecular weight of recombinant P-secretase was increased from 57 to 59 kDa. A lectin blot analysis indicated that recombinant beta-secretase from Tn5B1-4PSEC/GalT-ST cells (Tn5B1-4 cells co-transformed with cDNAs encoding beta-secretase, glycosyltransferases, GaIT, and ST) contained the glycan residues of beta 1,4-linked galactose and alpha 2,6-linked sialic acid. Two-dimensional electrophoresis revealed that recombinant beta-secretase from Tn5B1-4 beta SEC/GalT-ST cells had a lower isoelectric point than beta-secretase from control Tn5B1-4 beta SEC cells (Tn5B1-4 cells transformed only with beta-secretase cDNA). The enzyme activity of recombinant beta-secretase from Tn5B1-4 beta SEC/GalT-ST cells was enhanced up to 77% compared to control Tn5B1-4 beta SEC cells. The concentrations at half-maximum inhibition (IC50) values estimated from inhibition analyses using purified beta-secretases from Tn5B1-4/beta SEC and Tn5B1-4/beta SEC/GalT-ST cells were 32 and 290 nM, respectively. (c) 2005 Elsevier Inc. All rights reserved.