Functional expression in Aspergillus oryzae of p15, a protein with potent neurite-inducing activity in PC12 cells.
We previously reported that a fungal protein, p15, induces neurite outgrowth and differentiation of rat pheochromocytoma PC12 cells through the activation of the Ca2+ signaling pathway. We report here the secretory production of p15 in Aspergillus oryzae. Analysis of culture supernatant of A. oryzae transformed with the gene encoding the p15 precursor tagged with a hemagglutinin (HA) epitope demonstrated that the transformant secreted a protein with an apparent molecular mass of 17.5 kDa, which is a little larger than the expected size of mature p15-HA. By heat denaturation and ion exchange chromatography, p15-HA was easily purified from the culture supernatant with sufficient abundance. Although purified p15-HA was less active than the native p15 obtained from the culture broth of a producing fungal strain, it had neurite-inducing activity in PC12 cells in a dose-dependent manner, providing a system to study the action mechanism of p15.
Nakashima
Satoru
S
Department of Biotechnology, The University of Tokyo, Japan.
Arioka
Manabu
M
Kitamoto
Katsuhiko
K
D000818:Animals; D000903:Antibiotics, Antineoplastic / Q000096:biosynthesis* / Q000494:pharmacology*; D001236:Aspergillus oryzae / Q000378:metabolism*; D001483:Base Sequence; D002852:Chromatography, Ion Exchange; D004591:Electrophoresis, Polyacrylamide Gel; D000939:Epitopes / Q000276:immunology; D005656:Fungal Proteins / Q000096:biosynthesis* / Q000235:genetics*; D015966:Gene Expression Regulation, Fungal / Q000235:genetics; D006388:Hemagglutinins / Q000737:chemistry; D008969:Molecular Sequence Data; D016501:Neurites / Q000502:physiology* / Q000648:ultrastructure; D016716:PC12 Cells; D010957:Plasmids / Q000235:genetics; D051381:Rats
D000903:Antibiotics, Antineoplastic; D000939:Epitopes; D005656:Fungal Proteins; D006388:Hemagglutinins