EXPRESSION OF CELL-SURFACE GPI-ANCHORED HUMAN P97 IN BACULOVIRUS-INFECTED INSECT CELLS

The baculovirus/insect cell system (Autographa californica multiple nu clear polyhedrosis virus/Spodoptera frugiperda Sf9 cells) was used to express the GPI-anchored human melanomatumor antigen, melanotransferri n or p97. This system served to study the expression and productivity of recombinant GPI-anchored p97 by insect cells. The Sf9 cells express ed a cell surface GPI-anchored form of p97 as well as a soluble form o f p97 that did not appear to be derived from the GPI-anchored form of p97; Both recombinant forms, although Endo H resistant, migrated sligh tly faster (similar to 88 kDa) than the native p97 (similar to 95-97 k Da). The insect GPI-anchored p97 was sensitive to PI-PLC, which expose d a detectable crossreacting determinant. The Sf9 cell surface p97 exp ression was similar to that of human melanoma (SK-MEL-28) cells, where as the Sf9 cell specific secretion rate was 10-fold higher. Also Sf9 c ells retained considerably higher levels of p97 within the cell. The S f9 cell surface expression of p97 varied with time after infection, wi th the maximum expression, which appeared independent of multiplicitie s of infection greater than 1, occurring at 48 h. After 48 h, levels o f cell surface and secreted p97 fell whereas p97 retained within the c ell increased, which possibly reflected the lytic nature of the expres sion system. The successful expression of GPI-anchored human p97 by th e baculovirus/insect cell system not only provides a source of p97 for further research but also is the basis of an alternative method for t he commercial production of GPI-anchored proteins. (C) 1997 John Wiley & Sons, Inc.