DISRUPTION OF THE SACCHAROMYCES-CEREVISIAE YAP3 GENE REDUCES THE PROTEOLYTIC DEGRADATION OF SECRETED RECOMBINANT HUMAN ALBUMIN

Expression of recombinant human albumin (rHA) in Saccharomyces cerevis iae resulted in secretion of both mature albumin and a 45 kDa degradat ion product, comprising an N-terminal fragment of rHA with heterogeneo us C-termini between residues 403 and 409 (Geisow et al., 1991). Site- directed mutagenesis of the human albumin gene (HA) to change Arg(410) to Ala (R410A) caused a significant reduction in the amount of fragme nt produced. Mutation of the adjacent dibasic site Lys(413) Lys(414) h ad little effect in isolation, but in combination with the R410A mutat ion resulted in a further reduction in the amount of rHA fragment prod uced. This reduction could be duplicated with nature-identical rHA by disruption of the gene for an aspartyl protease (YAP3), alone or in co njunction with disruption of the KEX2 gene. Disruption of KEX2 alone d id not result in any improvement in the degree of degradation of the r HA. Reduced degradation was also observed when an rHA-human growth hor mone fusion protein was secreted from a yap3 strain, suggesting that s uch strains may have a general utility for heterologous protein secret ion. (C) 1998 John Wiley & Sons, Ltd.