High-level secretion of a Rhizopus niveus aspartic proteinase in Saccharomyces cerevisiae.

The gene encoding an extracellular Rhizopus niveus aspartic proteinase I (RNAP-I) was introduced into Saccharomyces cerevisiae. The yeast cell carrying a plasmid containing the intact RNAP-I gene under the control of the glyceraldehyde 3-phosphate dehydrogenase (GAPDH) gene promoter of S. cerevisiae did not synthesize RNAP-I at all. On the other hand, when the intron of the RNAP-I gene had been removed from the gene in the plasmid, the cell secreted RNAP-I with high efficiency. Processing of the pro-sequence occurred at the same region of the pro-enzyme during cultivation as observed in the culture of R. niveus. Moreover, the promoter and the terminator of the original RNAP-I gene were found to be weakly functional in the yeast cell with respect to expression of the intronless RNAP-I gene, although the initiation and termination sites were heterogeneous. The effects of vector-types on the extracellular production of RNAP-I were also investigated.