HIGH-LEVEL PRODUCTION OF HUMAN PARATHYROID-HORMONE (HPTH) BY INDUCED EXPRESSION IN SACCHAROMYCES-CEREVISIAE

Saccharomyces cerevisiae was used as host for high-level production of intact human parathyroid hormone (hPTH). The yield increased about 30 -fold by changing from the constitutive MF alpha promoter to the induc ible CUP1 promoter in the expression cassettes, use of another host st rain, and optimization of growth conditions where especially the pH va lue was crucial. The secreted products consisted mainly of intact horm one, hPTH(1-84). In addition, two C-terminally truncated forms that la cked the four or five last amino acid residues, hPTH(1-80) and hPTH(1- 79), were identified. These hPTH forms migrated aberrantly by SDS-PAGE as 14-kDa proteins, while the real masses measured by mass spectromet ry on HPLC-purified products were about 9 kDa. Availability of such ea sily purified truncated forms will be valuable for studies of how the C-terminal residues affect the structure and function of the hormone. Combination of mutations and disruptions of the host genes encoding pr oteinase A, B, carboxypeptidase Y, and Kex1p or Mkc7p did not influenc e the C-terminal deletions. The secretion of hPTH could be enhanced by overexpression of the yeast syntaxin gene SSO2, but the total level o f the hormone was not improved due to impaired growth. (C) 1998 Academ ic Press.